The Effects of Intraruminal Infusions of Sucrose and Xylose on Nitrogen And

JOURNAL OF AGRICULTURAL SCIENCE IN FINLAND Maataloustieteellinen Aikakauskirja Vol. 59: 405—424, 1987

The effects of intraruminal infusions of sucrose and xylose on nitrogen and fibre digestion in the rumen and intestines of cattle receiving diets of grass silage and barley

PEKKA HUHTANEN Department of Animal Husbandry, University of Helsinki, SF-00710 Helsinki, Finland

Abstract. Two male cattle(live weight 240 kg) fitted with rumen and simple T-piece duodenal cannulas were given a basal diet of 12 kg of silage (227 g/kg DM, 25.7 g N/kg DM), 1 kg of rolled barley and 100 g of mineral mixture. In addition to the basal diet, 0, 450 or 900 g/d of either sucrose or xylose was continuously infused intraruminally.

The amount of organic matter (OM) entering the duodenum (P < 0.001) and excreted

in the faeces (P < 0.01) was linearly increased with the increased sugar infusion. The proportion of digestible OM apparently digested in the rumen averaged 0.694, and did not differ (P > 0.05) with the level or type of sugar. Rumen ammonia concentration and molar proportions of isovalerate were decreased (P < 0.001) with increased sugar level. The decrease in the ammonia concentration was slightly

less (P < 0.05) with xylose than with sucrose. Molar proportions of acetate, propionate and butyrate were not affected by the sugar level but sucrose produced a lower (P < 0.05) proportion of acetate and higher (P < 0.01) proportion of butyrate than did xylose. There was a net loss of N (11.6 g/d) between the mouth and duodenum when the basal diet was consumed alone but a net gain (21.6 g/d) with the high level of sugar infusion. Micro- bial N flow at the duodenum, measured on the basis of RNA purine bases, increased linearly (P <0.001) with sugar level. The efficiency of microbial N synthesis (g N/kg OM apparently digested in the rumen) was 24.0, 29.1 and 30.0 (P > 0.05) for sugar levels of 0, 450 and 900 g/d, respectively. The amount of microbial N produced was closely related to rumen ammonia concentration (r —0.86; P < 0.001) and outflow of water at the duodenum (r0.83; P <0.001). The amounts of neutral detergent fibre (NDF) and acid detergent fibre (ADF) flowing

at the duodenum (P < 0.01; P < 0.001) and excreted in the faeces (P < 0.05) were increased linearly with sugar level. The increased amounts of fibre digested in the large intestine with sugar infusions were insufficient to compensate for the reduced digestion in the rumen and, as a result, overall digestibilities ofNDF and ADF were lower (P < 0.05). The effect of sugar level on the disappearance of silage N-free OM was consistent with the in vivo digestibility of fibre, although the extent of reduction seemed to be less in nylon bag incubations.

Index words: sucrose, xylose, silage, microbial synthesis, digestion

405 Introduction The different rates of energy and protein release suggest that silages should be supple- The change that occurs in nitrogenous com- mented with readily fermentable carbohydrates ponents of grass during ensiling is often ex- to promote microbial protein production. No tensive and varies with the type of fermenta- significant improvements in the efficiency of tion that occurs (McDonald 1982). Depending microbial protein synthesis have been found on the method of silage making, a substan- when barley supplements have been given tial proportion of the silage nitrogen (N) may (Thomas et al. 1980a; Rooke et al. 1985a). N be present as non-protein nitrogenous con- retention has frequently been improved with stituents (NPN). barley supplements (Thomson 1968; Kelly The efficiency of microbial protein synthe- and Thomas 1978), but this effect was not sis in the rumen in animals given silage diets noted by Kaiser et al. (1983) for maize has been reported to be lower than in animals starch. The effect of barley and of starch given hay or mixed diets. The Agricultural supplements on rumen ammonia concentra- Research Council (1984) reported a value of tion has been varied and sometimes no reduc- 23 g N/kg organic matter apparently digested tion has been found (Syrjälä 1972; Thomas in the rumen (Omadr) for the efficiency of et al., 1980a; Kaiser et a!., 1983; Chamber- microbial protein synthesis for silages as com- lain et al. 1985). The benefits of increased pared to 32 g N for hays and grasses. Thomas energy supply are partly offset by the in- and Chamberlain (1982) adopted a mean creased number of protozoa and intraruminal value of 25 N/kg Omadr for silage diets. The recycling of N (Chamberlain et al. 1985). reason for the low efficiency of microbial Sugar supplements have been shown to be synthesis is not entirely understood, but in more effective than starch supplements in part it may be due to the presence of silage reducing rumen ammonia concentration fermentation products and hence to the lower (Syrjälä 1972, Chamberlain et al. 1985), yield of ATP/kg Omadr. Extensively fer- and xylose more effective than sucrose mented silage may contain up to 150 g lactic (Chamberlain et al. 1985). Unlike starch, acid/kg dry matter (DM), and together with sugars have not increased the rumen protozoal the volatile fatty acids may account for 0.26 number (Chamberlain et al. 1985). of the metabolizable energy (Miller 1982). It The purpose of the present study was to has been suggested that one reason for low quantify the effects of sugar supplements on efficiency of microbial synthesis is poorly rumen microbial protein production in cattle matched rates of ammonia and energy release given a basal diet of grass silage and barley. from silage. However, Thomas and Thomas To avoid the negative effects of sugar supple- (1985) concluded that with extensively fer- ments on rumen pH, sugars were given in the mented silage the effect is mediated via energy form of continuous intraruminalinfusions. As supply rather than proteolysis per se. Re- another part of the study, it was investigated ducing the proportion of NAN in silage from whether the adverse effect of readily fermen- 0.56 to 0.40 by increasing the application of table carbohydrates on rumen cellulolysis formic acid from 0 to 5.9 litres/t had little could be alleviated by the continuous supply influence on rumen ammonia concentration, of supplements that would reduce the post- bacterial N synthesis or passage of undegraded prandial change in rumen pH. dietary protein to the small intestine (Cham- berlain et al. 1982). Rooke et al. (1985a) Materials and methods suggested that the nature of the nitrogenous substances available to rumen microbes may Animals limit the efficiency of microbial synthesis with The experimental animals were two silage diets. Ayrshire male cattle, aged 8 months at the 406 time of surgery and weighing 240 kg at the with 900 g of sucrose for days 17—24. At the beginning of the experiment. The animals same time animal B was infused with water, were fitted with a permanent rumen cannula 450 g of xylose and 900 g of xylose, succes- and a simple T-cannula in the proximal duo- sively. In the second period the infusions of denum, approximately 5 cm posterior to the the animals were reversed. After the first pe- pylorus and proximal to the point of entry of riod about 10 kg of rumen contents from a bull the common bile duct. The animals were held given a similar basal diet was transferred into in metabolism crates. Feed was given in two the rumen of each animal to hasten the adap- equal meals at 9 hours and 21 hours. Water tation to the new diet. The animals were then was freely available. given a rest period of 5 d before the second period commenced. The sugars was infused at a rate of 75 ml/h with a peristaltic pump Feeds (Watson —Marlow Ltd, Cornwall). The basal diet consisted of 12 kg of grass Cr-labelled straw was used as particulate silage, 1 kg of rolled barley (240 g/kg of total marker and Coedta as liquid phase marker. dry matter (DM) intake) and 100 g of com- Markers were prepared as described by Uden mercial mineral mixture. Silage was from et al. (1980). Fifteen grammes (2 X 7.5 g) of second cut timothy, meadow fescue and clover Cr-straw (68 mg Cr/g) per day was adminis- sward, harvested with a precision-chop forage trated via rumen cannula at feeding times and harvester and ensiled with of a solution of 4 grammes of Coedta per day was infused formic acid (800 g formic acid/1) at a rate of continuously with the nutrient infusates. The 5 1/t. The silage was the same as that used in amount of constituents excreted in faeces was the dairy cow experiment (Huhtanen 1987). determined using Cr as an indigestible marker. Both animals were also infused intraruminally Faeces were collected totally on day 8 of each with three different solutions, succesively. The subperiod and mixed thoroughly, and a chemical composition of the dietary ingre- representative sample was taken. dients is given in Table 1. Duodenal samples (approximately 120 ml) were taken on day 8 of each subperiod at 2 h intervals through a 24 h period and pooled to Experimental procedure provide a sample. This sample was further The experiment consisted of two periods of divided into two parts, of which one part was 24 days, with each divided into three sub- further divided by centrifugation (800 g for periods of 8 d. Intraruminal infusions of 10 min) into liquid phase and particulate sucrose and xylose were given at three dif- matter. The unrepresentative digesta sample ferent levels: 0 (control), 450 (low) and 900 and the particulate matter were oven-dried at (high) g/d. During the first period animal A was 60 °C, milled and stored before analyses. The intraruminally infused with water for days liquid phase was stored at 0 °C before the I—B,1 —8, with 450 g of sucrose for days 9—16 and analysis of markers. Rumen samples were

Table I. Chemical composition of experimental feeds.

In dry matter (g/kg) DM (g/kg) Ash N NDF ADF C HC ADL

Silage 227 89 25.7 582 346 313 236 33 Barley 867 28 19.6 228 76 62 152 14

C = cellulose, HC = heraicellulose, ADL = acid detergent lignin. In silage: pH 4.14; in DM (g/kg); sugars 26,

lactic acid 41, acetic acid 18, propionic acid 0.3, butyric acid 0.3; in total N (g/kg): NH 3 N 50, soluble N 460. 407 taken before feeding and thereafter 1,2, 3, determined by the method of Zinn and 4,6, 8 and 10 h after feeding. Owens (1982). The concentrations of Cr and The rate of disappearance of dry matter Co in digesta and faecal samples were deter- (DM), organic matter (OM), N-free OM, re- mined by atomic absorption spectrophoto- sidual fraction and nitrogen (N) of the silage metry after ashing of the samples and diges- used in the experiment and of the untreated tion with a potassium bromate acid mixture barley straw was determined in the rumen of (Williams et at. 1962). both animals for each infusion. The residual fraction was calculated as OM minus crude Calculation of results protein, sugars and fermentation acids. Fer- mentation acids and sugars were assumed to The flow of digesta DM and other con- be lost in washing and were not determined stituents entering the small intestine was cal- from the residues. From day 6 to day 8 of each culated by Faichney’s (1975) double marker subperiod, bags (porosity 40 /un) containing method, with the modification that marker 3.5 g of DM were incubated for periods of 12, concentrations in the unrepresentative digesta 24 and 48 (9 bags/feed, 18 bags/animal) and sample and solid phase were used to calculate a ’0 h wash’ value was determined for three the reconstitution factor (France and Siddons bags per feed. 1986). The flow of N in the microbial fraction at the duodenum was calculated by re- ference to purine in digesta. The value 0.15 Analytical methods for the ratio nucleic acid (RNA-basis) N:total Oven DM content of silage was corrected N in rumen bacteria was used to calculate the for volatile losses of lactic acid, volatile fatty flow of microbial N (ZiNNand Owens 1986). acids (VFA) and ammonia as described by Feed N degradability was calculated as (1 Porter et at. (1984). Fermentation quality of (Total N ammonia N microbial N the silage was analysed by the methods endogenous N))/Feed N. A value of 130 mg/ 0 75 described by Fluhtanen (1987). N content of kg - was used to estimate the flow of feeds, digesta samples and faeces was deter- endogenous N into the duodenum (Orskov mined by Kjeldahl method. Neutral detergent and MacLEOD 1983). Outflow of water was fibre (NDF), acid detergent fibre (ADF) and calculated as total digesta flow minus DM acid detergent lignin (ADL) were analysed ac- flow. cording to GoEßiNcand Van Soest(l97o), but as modified by Robertson and Van Soest Statistical analyses (1977) for barley samples. Hemicellulose was The statistical model used to analyse the calculated as the difference between NDF and flow data of different constituents was ADF, and cellulose as the differencebetween + + d) ADF and ADL. In vitro digestibility of silage y,jkim -n +Aj+Pj Lk +S, eiJk/m was determined by the two-stage procedure of where A, P, L and S are the effects of animal, Tilley and Terry (1963) to control the varia- period, sugar level and sugar. The effect of tion in silage quality during the experiment. the sugar level was further partioned into the The ammonia N concentration in rumen effect due to linear and quadratic trends samples and in the liquid phase of duodenal (Snedecor and Cochran 1967). A split-plot digesta was determined by the method of analysis of variance was fitted to the data McCullough (1967). VFAs in rumen fluid from rumen fluid analyses: samples taken before feeding and 2, 4 and 6 h

+ + + + + after feeding were determined by gas liquid Vijklnm —H +Ai Pj Lk +Sl eUk, H„, + + + + chromatography (Huida 1973). Purine con- (AH)im (PH) jm (LH)km (SH),m p tents of digesta samples and faeces were ijklmn (2)

408 effect NS NS NS NS NS NS NS NS of Level Quadratic

*** significance Linear *» ** ** NS NS NS NS

Sugar NS NS NS NS NS NS NS NS barley. Statistical and silage L 78 49 40 32 0.010 0.016 0.016 0.013 grass SEM of S 0.008 0.013 0.013 0.011 diet 64 40 32 26 given cattle Xyl 0.750 0.692 0.307 0.479 in 4276 2222 2054 1067 1067 0.001) 900 < digestion (P Sue 0.751 0.681 0.318 0.487 *** (OM) 4276 2191 2085 1065 1065 0.01),

< matter level (P Xyl 0.748 0.696 0.304 0.476 ** organic Sugar 3807 1984 1824 867 956 on 450 0.05), < 0.656 0.344 0.501 (P 0.745 * infusions Sue 3807 1860 1947 977 970 (Xyl)

xylose 0 0.739 0.705 0.295 0.455 3316 1729 1587 723 864 (non-significant), and NS (Sue) sucrose Significance: of h) rumen and (g/24 the large level. duodenum = effect of intestine in duodenum small The in digestibility matter OM and small L matter small proximal in intestine 2. proximal intestines large sugar, food Digested faeces organic small at large = Organic In Fermented At In Apparent Disappearance digestible Before In intestines digested Table of OM and S 409 where A, P, L and S are the same effects as (P < 0.001) increased with sugar level. On the in model (1), eukl is the main plot error and average, microbial N production was in- H is the effect of sampling time. creased by 25 g/kg sugar infused. The quan- tity of N excreted in faeces was increased with

sugar level (P < 0.01) and the resulting fall Results in apparent digestibility of N was significant

(P < 0.05). On the other hand, N entering the Digestion of organic matter duodenum was more digestible (P < 0.05) DM intake of the basal diet was 60 g DM/kg when sugars were infused.

W 0 75 . The amounts of ingested organic The efficiency of microbial N synthesis (g matter (OM) and the amounts of OM N/kg OM apparently digested in the rumen measured at the proximal duodenum and in (Omadr)) averaged 27.8 g N/kg Omadr, and the faeces are shown in Table 2. The amounts was not significantly affected either by the of OM entering the proximal duodenum and level or the type of sugar. When expressed in which were subsequently voided in faeces were relation to OM truly digested in the rumen, linearly increased (P < 0.001; P < 0.01) with by assuming that microbial matter had a sugar level. Differences between the sugars in N:OM ratio of 0.09 (Czerkawski 1986), respect of digestion of OM were small and values showed a narrower range with a mean non-significant. The amount of OM apparently value of 21.2. However, there was a trend digested in the rumen was increased (P < toward higher efficiency when sugars were 0.01) with sugar level. There was a tendency given, and more so for sucrose than xylose. for the total OM digestibility to be higher On basal diet there was a substantial loss of when sugar supplements were given. The N between the mouth and duodenum, but average increase in digestible OM intake was with sugar infusions the flow of NAN at the 0.84 kg/kg sugar infused. duodenum exceeded the amount of N ingested The proportion of digestible OM disap- by 0.07—0.23. Outflow of water at the duo- pearing in the rumen averaged 0.694, and was denum was increased (P < 0.05) with sugar not affected either by the type or level of infusions. sugar. The amount of OM digested in the The apparent loss of RNA between duo- small and large intestines was linearly (P < denum and faeces was 0.818 (SE 0.010). 0.01) increased with sugar level. The propor- Faecal N loss was closely related to the passage tion of OM entering the duodenum that of NAN at the proximal duodenum: (Faecal

= disappeared in the intestines averaged 0.467, N (g) = 0.155* NAN + 10.5; r 0.80**). and tended to be higher when sugar supple- Based on this equation, the true digestibility ments were given. of NAN was 0.845 and metabolic faecal N 10.5 g, which is equivalent to 2.5 g N/kg DM intake. N metabolism

The average N concentration in the diet was Digestibility offibre components 24.3, 21.1 and 19.7 g/kg DM for infusion levels of 0, 450 and 900 g, respectively. There NDF intake ranged from 1755 to 1799 g/d was a linear (P < 0.01) increase in the flow and intake of ADF from 986 to 1011 g/d of total N at the duodenum with sugar level. (Table 4). Increasing the amount of sugar Ammonia N flow averaged 3.1 g/d and was infused resulted in a linear increase in the flow unaffected by the supplementation. The pat- of NDF (P < 0.01) and ADF (P < 0.001) at tern of flow of non-ammonia N (NAN) was the proximal duodenum and a fall in the quan- therefore similar to that of total N (P < tity of NDF and ADF fermented in the rumen

0.01). The flow of microbial N was linearly (P < 0.05, P < 0.01). With sugar diets the

410 (1/day) effect NS NS NS NS NS NS NS * NS NS NS NS NS water of Level Quadratic of *** ** ** ** * * ** * NS NS NS NS NS 5. significance Linear 07 outflow W and Statistical Sugar NS NS NS NS NS NS NS NS NS NS NS NS NS mg/kg 130 nitrogen of of L 0.78 4.7 0.14 4.7 2.5 1.1 0.013 0.007 0.056 2.20 1.27 0.036 2.9 secretion digestion SEM N tract, S 0.63 3.8 0.11 3.8 2.1 0.9 0.010 0.006 0.046 1.80 1.04 0.029 2.4 digestive endogenous the Xyl 3.2 62.6 27.0 0.704 0.750 1.17 28.4 21.6 0.604 59.5 Assuming 91.7 110.4 107.2 ' through 900 1.23 rumen; nitrogen Sue 91.7 116.2 2.8 113.5 67.9 30.2 0.672 0.739 31.6 23.3 0.595 60.8 the of in flow digested and level 0.001) Xyl 88.0 96.4 3.2 93.1 52.5 24.1 0.724 0.749 1.07 26.5 20.5 0.630 52.6 truly < barley. Sugar (P intake 450 *** on and matter Sue 88.0 105.2 3.0 102.3 58.7 25.3 0.711 0.760 1.16 31.6 23.5 0.596 59.1 0.01), silage Organic < infusions (P grass 2 (Xyl) ** of rumen; 0.743xylose diet 0 3.2 22.0 0.753 0.715 0.83 19.2 0.05), 89.2 77.6 74.3 42.0 24.4 45.4 the < and given in (P (Sue) cattle digested * in

N N 1 2 sucrose of of Nof N (1/day) apparently (non-significant), h) feed duodenum N N OMADR OMTDR effect food duodenum duodenum/ of NS in digestibility digestibility N/kg N/kg water The from (g/24 N matter N of 3. duodenum entering proximal Total Ammonia Microbial faeces Nitrogen Total At Non-ammonia In entering intake Degradability Organic Significance: Apparent Apparent 1 Table NAN N Microbial Microbial Outflow 3 411 effect barley. of NS NS NS NS NS NS NS NS NS NS NS NS NS NS and Level Quadratic *•• ** * * * ** * * silage significance Linear NS * * NS NS NS grass of Statistical Sugar NS NS NS NS NS NS NS NS NS NS NS NS NS NS diet given L 0.014 0.014 cattle 19 46 31 32 0.021 10 20 12 19 0.022 0.017 0.017 in SEM S 0.017 0.011 0.011 digestion 16 37 26 26 8 17 10 15 0.018 0.014 0.014 (ADF)

Xyl 0.591 0.948 0.052 0.596 0.925 detergent 1774 996 778 723 986 542 443 397 0.075 acid 900 and Sue 0.576 0.888 0.112 0.572 0.931 0.069 (NDF) 1774 912 861 750 986 525 459 419 level 0.001) detergent Xyl 0.631 0.937 0.063 0.635 0.924 0.076 < Sugar 1799 1066 733 663 1011 593 418 369 (P neutral 450 *** on Sue 758 689 0.617 0.940 0.060 597 414 387 0.617 0.959 0.041 0.01), 1799 1041 1011 < (P infusions ** (Xyl) 0 0.05), 596 582 0.668 0.988 0.012 988 346 334 0.676 0.982 0.018 xylose 1755 1158 642 < (P and * (Sue) sucrose rumen digestible rumen digestible intestines (non-significant), of the large the large of intestine of intestine in duodenum in duodenum NS Effect h) digestibility small and h) digestibility small and 4. (g/24 food proximal faeces small (g/24 food proximal faeces small In Fermented At In Apparent NDF Disappearance Before In intestines In Fermented At In Apparent ADF Disappearance ADF Before In Significance: Table NDF of NDF ADF of 412 effect NS NS NS NS NS NS NS NS NS NS NS NS NS NS of Level Quadratic *** * ** * * * ** significance Linear NS • NS * * NS NS barley. Statistical Sugar NS NS NS NS NS NS NS NS NS NS NS NS NS NS and silage L 11 17 10 13 0.016 0.007 0.007 9 29 22 13 0.019 0.035 0.035 grass SEM of S diet 9 14 8 11 0.013 0.006 0.006 7 24 18 11 0.016 0.028 0.028 given cattle in Xyl 901 568 333 280 0.688 0.915 0.085 788 453 335 326 0.586 0.979 0.021 digestion 900 Sue 901 561 339 298 0.668 0.931 0.069 788 387 401 330 0.581 0.840 0.160 hemicellulose 0.001) and level Xyl 0.727 0.908 0.092 0.627 0.951 0.049 < Sugar 929 614 316 253 788 473 316 293 (P cellulose 450 •*• on 0.01), Sue 929 621 309 268 0.711 0.940 0.060 788 344 302 0.616 0.916 0.084 444 < (P infusions ** (Xyl) 0 0.749 0.952 0.048 0.684 0.996 0.004 0.05), xylose 904 649 255 227 767 516 251 248 < (P and * (Sue) sucrose rumen digestible h) rumen digestible intestines (non-significant), of h) the large large of intestine (g/24 the of intestine in duodenum in duodenum NS Effect (g/24 digestibility small and digestibility small and 5. food proximal faeces small food proximal faeces small In Fermented At In Apparent cellulose Disappearance Before In intestines In At In Apparent hemicellulose Disappearance Before In Significance: Table Cellulose of cellulose Hemicellulose Fermented of hemicellulose 413 amount of NDF digested postruminally in- overall digestibility of cellulose from 0.749 to creased from 14 (control) to 70 (low) and 0.678 and of hemicellulose from 0.684 to 83 g/h (high). However, increased NDF and 0.584. The proportion of digestible cellulose

ADF digestion in the intestines was insuffi- fermented in the rumen was decreased (P < cient to compensate for the reduced fibre 0.05) due to the sugar infusions and the same digestion in the rumen. There was a linear tendency was found for hemicellulose. increase (P < 0.05) in faecal output NDF and ADF with sugar level. Total digestibilities of Rate of degradation of silage NDF and ADF were higher (P < 0.05) when the basal diet was given alone than when supplemented with the high level of sugar Values describing the disappearence of (0.668 vs. 0.584; 0.676 vs. 0.584). With both silage constituents from nylon bags incubated levels of xylose, NDF and ADF digestibilities in the rumen are presented in Table 6. The were marginally higher than they were with disappearance of silage OM, N-free OM and sucrose. The proportion of NDF digestion residual fraction was reduced with increased occurring in the rumen fell linearly (P <0.05) level of sugar infusion, but only after incu- with sugar level; a similar tendency was found bation of 48 h were the linear effects of sugar for ADF, but the difference was not signifi- level significant (P < 0.01, P < 0.001). After cant. an incubation period of 24 h the difference

Values for cellulose and hemicellulose was close to becoming significant (P = 0.066). digestion indicate similar effects due to the Because sugar infusions tended to increase N level of sugar infusion as observed for NDF loss, the disappearance of N-free OM was and ADF (Table 5). Increasing the level of more strongly affected by the sugar infusions sugar decreased the quantities of cellulose and than the disappearance of DM or OM. The hemicellulose fermented in the rumen, with a content of the residual fraction in silage was consequent linear (P < 0.05) decrease in the 667 g/kgDM, which is slightly higher than the

Table 6. Effect of sucrose (Sue) and xylose (Xyl) infusions on the disappearance(mg/g) of different constitutents of silage from nylon bags incubated in the rumen of cattle.

Sugar I cvcl SEM Statistical significance of effect W ~0 «Ö W ~s" r Sugar Level'

OM 12 h 420 447 460 412 428 15.4 18.8 NS NS 24 h 599 603 616 598 587 8.9 10.9 NS NS 48 h 699 707 717 701 691 4.6 5.7 NS •• Nitrogen 12 h 741 753 753 730 758 12.4 15.2 NS NS 24 h 850 850 845 843 862 5.0 6.1 NS NS 48 h 877 882 875 876 887 4.4 5.3 NS NS N-free Om 12 h 351 382 398 345 357 16.5 20.2 NS NS 24 h 545 550 568 546 529 10.0 12.3 NS NS 48 h 664 670 684 663 649 5.0 6.1 NS •*• Resid. fract. 2 12 h 268 302 320 260 274 18.7 22.9 NS NS 24 h 486 492 517 487 468 11.2 13.8 NS NS 48 h 617 627 643 620 604 5.6 6.9 NS ••

2 S = sugar; L = level; 1 Linear trend of sugar level, no significant quadratic trend; OM minus crude protein, sugars and fermentation acids. 414 for content of NDF (582 g/kg DM), and thus effect changes in degradation of the residual frac- of NS * NS NS NS NS * * NS Values Quadratic tion may reflect changes in NDF degradation. Level No significant differences between the sugars *** *** were found, barley. significance Linear NS NS NS NS NS * NS although disappearances of OM and and N-free OM were marginally higher with the xylose infusion. The '0 h wash' value (mg/ •• silage Statistical Sugar NS * NS * NS NS * * g) was 186 for OM, 512 for N 122 for N-free grass OM 122 and 4 for the residual fraction. The average disappearance of straw DM of was 6.1 °Io lower for the high level of sugar diet L 0.023 0.34 3.13 6.9 3.5 4.6 0.32 0.29 0.64 than for the basal diet after incubations of 24 given and 48 h. SEM cattle S in 0.018 0.27 2.56 5.6 3.5 3.8 0.26 0.23 0.53 Rumen fermentation (VFA) Mean values of rumen pH, concentrations of ammonia N and total VFA and the molar acids proportions of individual fatty acids are given Xyl 6.56 5.37 9.9 fatty 93.9 684 186 104 11.5 4.6 in Table 7. Rumen ammonia N concentration and molar proportion of isovalerate were 900 markedly reduced when the of sugar times. amount volatile Sue 6.57 4.26 9.6 7.7 in the basal diet was increased. There were and 94.2 665 174 132 12.2 significant linear (P < 0.001) and quadratic N sampling (P < 0.05) trends. The effect of sugar level 4 on the molar proportion of valerate was 3 of NH level 0.001) significantly linear (P < 0.05) and quadratic pH, VFA Xyl 6.65 5.91 96.9 691 172 110 11.7 10.8 5.4 < for Sugar (P and 450 *** rumenon times Sue 6.64 5.05 94.4 656 179 133 11.8 12.2 7.0 0.01), < (P infusions sampling ** (Xyl) 8 0 0.05), of 6.63 8.30 97.6 175 115 16.3 13.1 7.4 xylose 675 < (P and averages * are (Sue) N sucrose NHj (non-significant), of and (mmol/1) acid acid NS Effect pH (mmol/1) (mmol/mol) acid acid acid N proportions acid Fig. I. The effect of sugar infusions on rumen ammo- 7. VFA nia concentration in cattle given diets of grass 3 Propionic Butyric VFA Acetic Valeric Caproic Significance: «ihge and barley (• 0 g/d; ■ 450 g/d; pH Isovaleric Table NH Total Molar of A 900 g/d). 415 Fig. 2. The effect of sugar infusions on rumen pH, total VFA and the proportions of individual fatty acids in cattle given diets of grass silage and barley (• 0 g/d; ■ 450 g/d; A 900 g/d).

416 (P < 0.05). Sucrose infusion resulted in A similar small increase in the total OM slightly, although significantly (P < 0.05), digestibility of the diet was observed by lower ammonia concentration than did xylose. England and Gill (1985) when increasing Concentration of total VFA and the molar levels of sucrose supplements were given with proportions of acetate, propionate and silage to young cattle. The increased amount butyrate were not affected by the level of of faecal OM with sugar infusions consisted sugar infusion. As compared to sucrose infu- mainly of NDF. The amounts of faecal OM sion, xylose infusion showed a significant not analysed as NDF were 282, 287 and (P < 0.05) increase in the proportion of 329 g/d for control, low and high sugar infu- acetate and an associated decrease (P < 0.01) sions, respectively. In vitro digestibility of in that of butyrate without any effect on the silage OM was 0.643, 0.646 and 0.650 when proportion of propionate. the sugar levels of 0, 450 and 900 g/d were With the sugar infusions, rumen ammonia infused and therefore, it can be assumed that concentration was below 3 mM before feeding the results were not affected by the variation and again fell below this level 6 h post-feeding in silage digestibility. Further, similar or (Fig. 1). The postprandial changes in am- higher increases in the output of faecal OM monia N concentration were significantly with increasing addition of sugar were

(P < 0.01) different between the basal diet obtained when Coedta or acid insoluble ash and diets with sugar infusions. (Van Keulen and Young 1977) was used as Postprandial changes in rumen pH, the con- indigestible marker. centration of total VFA and the proportions Similar values as in the present study for the of most of the individual fatty acids were proportion of OM entering the duodenum that smaller in the case of sugar infusions (Fig. 2). disappeared in the intestines can be calculated Interaction between the level of sugar infusion from the results of Thomas et al. (1980b) and and sampling time was significant as regards Rooke et al. (1985b) for sheep and cattle pH (P < 0.001), total VFA (P < 0.01), and given silage diets. The increased amount of the proportions of acetate (P < 0.01), pro- OM digested in the intestines when sugar was pionate (P < 0.01) and valerate (P < 0.05). infused can be attributed in part to increased The most pronounced differences between the synthesis of microbial matter in the rumen and basal diet and sugar infusions in the postpran- its subsequent digestion and in part to dial changes in rumen fermentation characte- increased digestion of NDF in the lower tract. ristics were in the quadratic effect of sampling time. No interactions between the type of N metabolism sugar and sampling time were found. The lower ammonia concentration in the rumen with sugar infusions is consistent with Discussion earlier observations of Syrjälä (1972) and Chamberlain et al. (1985). The effect of OM digestion sugar level on the ammonia concentration was The average value of 0.694 obtained for curvlinear. In contrast to the findings of digestible OM disappearing apparently in the Chamberlain et al. (1985), sucrose infusion rumen is in agreement with the value of 0.70 here produced a larger reduction in rumen calculated by Thomas and Chamberlain ammonia concentration than did xylose. This (1982) for sheep given silage based diets. A discrepancy may be related to the different slightly higher value of 0.73—0.75 for cattle method by which the sugars were incorporated given silage diets was adopted from the in the ration: Chamberlain et al. gave the literature by Agricultural Research Council sugars via the rumen cannula at feeding time. (1984). The higher fermentation rate of sucrose than 417 of xylose (Sutton 1968) causes a larger reduc- characteristics and supplemented with barley. tion in rumen pH and higher ammonia con- Changes in rumen ammonia concentration centration, and they concluded that the dif- reflected well differences in the net incorpo- ferent action of the sugars may in part be ration of ammonia by the rumen microbes associated with this reduction of rumen pH. (Fig. 3). Slightly lower production of micro- In the present study, continuous infusion of bial N with xylose is consistent with the higher the sugars did not reduce rumen pH below 6.4 ammonia concentration when xylose infusion with sucrose or xylose (Fig. 2). was given. It is likely that the effect of the Relative to the basal diet of silage and sugars on microbial synthesis was related to barley sugar infusions increased markedly energy supply per se rather than to the production of microbialprotein. Efficiency of synchronization of ammonia production and microbial protein synthesis has been found energy release as was proposed by Syrjälä lower in animals given silage diets than in (1972). The release of ammonia from silage those given hay, as summarized by Thomas N compounds was rapid (Fig. 1), but con- (1982) and Agricultural Research Council tinuous infusion of sugars supported an even (1984). One probable reason for the lower supply of energy during the whole feeding efficiency with silage diets is that silage OM cycle. Supporting evidence against the syncho- consists of fermentation products and the ronization theory is provided by Chamber- yield of ATP/kg Omadr is low. Another lain et al. (1985) who found a considerable reason may be that the rates of ammonia and influence on the time-course carbohydrate energy release are poorly matched in silage. fermentation but no influence on the rumen In thepresent study, the silage was not exten- ammonia concentration when barley supple- sively fermented and had a low concentration ments were fed 2 h before, 1 h before and at of fermentation acids and low proportion of the same time as silage. ammonia and soluble N in total N. The method by which the sugars are given The positive effect of sugar infusion on may have some effect on microbial synthesis. microbial N production in the rumen indicates Large amounts of sugars given in a single meal that microbial synthesis was limited by energy cause a rapid reduction in rumen pH. Russel supply even with silage of good fermentation and Dombrowski (1980) found in continuous culture that growth yields of most bacteria were depressed at pH values lower than 6.0, cellulolytic bacteria being the most sensitive. The advantage of an even supply of energy for microbial synthesis has been demonstrated with continuous feeding and increased feeding frequency in sheep (AlAttar et al. 1976) and in dairy cows (Tamminga 1979). Low ammonia concentration (0.5—3.0 mM) with sugar infusions 6 —12 h after feeding did not seem to limit microbial synthesis. Such a low concentration of ammonia N in the rumen of cattle fed silage diets has been previously noted by Rooked/ al. (1985b) and Thomson et al. (1981). There has been considerable Fig. 3. The relation of rumen ammonia concentration and duodenal flow ofmicrobial N in cattle given controversy over the optimum ammonia con- diets of grass silage and barley supplemented centration to sustain maximum microbial with intraruminal infusions of sugars, y = 88 yields. Satter and Slyter (1974) found (± 6.7) 3.86 (±0.74) x; n 12; r 0.856; P < 0.001). 1.4—3.6 mM to be adequate for maximum 418 growth of rumen micro-organisms in vitro, with the ammonia concentration in the rumen and the general conclusion is that 3.5 mM is (r o.9l***). adequate with normal forage concentrate diets An increasing liquid phase dilution rate has (Miller 1982). A mean ammonia concentra- been associated with an increase in the effi- tion of 3.5 mM will assure the bacteria enough ciency of microbial growth (Harrison and ammonia even during the low point of the am- Mc-allan 1980). This is largely because the monia concentration cycle (Satter 1982). proportion of energy consumed for mainte- Many rumen bacteria have a very high affinity nance diminishes. The liquid dilution rate was for ammonia, and these organisms can attain not determined in the present study, but out- 95 % of their maximal growth rate in the flow of water at the duodenum was markedly presence of even 1.0 mM ammonia (Schaefer increased by sugar infusions and the dilution et al. 1980). rate was likely increased as well. There was Of the individualfatty acids, only the molar a close linear relationship between the outflow proportion of isovalerate showed a significant of water and microbial N production (Fig. 4). correlation (r —o.BB***) with the amount of Most of the rumen bacteria are found to be microbial N produced. Examination of the associated with feed particles (Hobson and biochemical pathways concerned shows that Wallace 1982) and the rate of passage of the amount of ATP generated per mole of feed particles may not always be related to hexose fermented in the rumen is not markedly liquid phase dilution (Mathers and Miller influenced by the VFA pattern produced 1981). Soluble sugars were available in the (Tamminga 1979; Czerkawski 1986). Branched- liquid phase during the whole feeding cycle, chain volatile fatty acids are required for the which may change the ratio of attached to free biosynthesis of branched-chain amino acids or bacteria. This shift would increase the dilu- higher fatty acids (Allison and Bryant 1963). tion rate of microbes from the rumen because The lower proportion of isovalerate with sugar the dilution rate of liquid is normally higher infusion indicates efficient incorporation of than that of particles. this acid by rumen microbes. The molar pro- The depressive effect of sugar infusions on portion of isovalerate was highly correlated the apparent N digestibility is consistent with earlier experiments when sucrose (Syrjälä 1972; Englandand Gill 1985) or starch (Kai- ser et al., 1983) supplements were given with grass silage. In contrast, Gill and Ulyatt (1977) did not observe any effect on N digestibility when a mixture of starch and sucrose was infused in the rumen of sheep given silage. The increased amount of N voided in the faeces when sugar was infused can partly be explained by the larger amount of indigestible residue of bacterial protein produced in the rumen. Another factor may be increased carbohydrate fermentation in the hindgut, leading to synthesis of microbial biomass there and its excretion in the faeces (Armstrong and Smithard 1979). Fig. 4. The relation of duodenal flow of water and microbial N in cattle given diets of grass silage Fibre digestion and barley supplemented with intraruminal infusions of sugars, y = —l.l (±11.8) + 1.03 Increase in the sugar level resulted in a (±0.20) x; n 12; r 0.833; P < 0.001). significant reduction in the digestibility of

5 419 fibre components and in the proportion of severe in the case of disappearance from nylon ruminal digestion. A reduction in fibre or cel- bags than of in vivo digestion. DM intake was lulose digestibility has been noted in many higher with the sugar supplements, which may studies (Thomas et al. 1980b; Kaiser et al. have reduced digesta retention time in the 1983; England and Gill 1985), following the rumen (Growum and Williams 1977) and supplementation of forages with readily thus contributed to fibre digestion. Calcula- fermentable carbohydrates. The proportion of tion of degradation constants using the equa- digestible cellulose broken down in the rumen tions of orskov and McDonald (1979) and (0.93) agrees with the values reported by McDonald (1981) shows the sugar infusions Beever et al. (1977) and Thomas et al. to have increased the lag timeand indigestible (1980b), but lower values have been reported fibre pool without any change in the rate of especially with pelleted forage (Thomson et digestion. The lag time (h), potentially al. 1972) or with high proportion of concen- degradable pool (mg/g) and rate of digestion trate in the diet (MacRAE and Armstrong of N-free OM were for the control diet 1.9, 1969). The proportion of digestible hemicel- 720 and 0.062 and for sugar the infusions lulose disappearing in the rumen was similar (mean value) 4.1, 687 and 0.066, respectively. to that of cellulose. The increased lag time is in agreement with The increase in the postruminal digestion of Mertens and Loften(l9Bo) who noted, with cellulose and hemicellulose with sugar infu- starch supplement, an increased lag time of sions was insufficient to compensate the fibre digestion in vitro. However, in their reduced digestion in the rumen. The amount study thepotential extent and the rate of fibre of NDF excreted in the faeces was increased digestion were not affected by starch addition. by 0.16 g/g sugar infused and that of ADF The optimal ammonia concentration by 0.08 g. Digesta retention time in caecum needed to support a maximal rate of fermen- and colon is quite short; values of s—lo5 —10 h tation may be higher than that suggested to were obtained from direct measurements be minimum for microbial growth. Mehrez (Grovum and Williams 1977) or can be cal- et al. 1977) observed an increase in the rate culated from reciprocal of k 2 (Hartnell and of breakdown of barley until the ammonia Satter 1979; Huhtanen 1987). Because of the concentration reached 17 mM, whereas short digesta retention time and quite small Ortega et al. (1979) were unable to show any pool of digestible fibre entering the caecum, increase in the rate of fermentation by in- complete compensation of reduced fibre creasing the rumen ammonia concentration digestion in the rumen cannot be expected. from 4.5 to 19.6 mM. orskov(l9B2) found Moreover, iffibre digestion in the hindgut is an ammonia concentration of 1.5 mM to characterized by a similar lag time as in the maximize the rate of digestion of alkali-treated rumen (Mertens 1977), the amount of fibre straw. On the basis of the latter two studies digested postruminally would be expected to it would seem that fibre digestion was not be further reduced. limited by low rumen ammonia concentration The effect of sugar infusion on the dis- in the present study. appearance of OM, N-free OM and the Rumen cellulolysis appears to be inhibited residual fraction from nylon bags incubated by low rumen pH (pH effect) and by the avail- in the rumen was consistent with the in vivo ability of readily fermentable carbohydrates digestibility of the various fibre components (carbohydrate effect) (Mould et al. 1983). in the rumen and in the total tract. Correla- Cellulolysis in the rumen is inhibited at pH tion between the amount of the residual frac- 6.0—6.2 (Stewart 1977; Mouldand orskov tion disappearing in 24 h and in vivo NDF 1983). Russel and Dombrowski (1980) found digestibility was o.97*** (n 6). The adverse the growth of pure cultures of cellulolytic bac- effect of the sugar infusions tended to be less teria to decrease sharply when the pH fell

420 below about 6.3. In the present study rumen Rumen fermentation pH was not affected by sugar infusion and Continous infusion of sugars caused a was throughout the day above the critical stable fermentation with less diurnal variation values cited above. From this observation it in rumen pH, total VFA and proportions of can be concluded that the depression in fibre most individual fatty acids. As was found by digestion was due to the availability of readily Chamberlain et al. (1985) in vivo and Sutton fermentable carbohydrates. This is in agree- (1968) in vitro, xylose increased the propor- ment with Mould et al. (1983), who found tion of acetate and reduced of butyrate. that reduction in DM degradation with that In contrast to the of orskov and molasses supplements could not be alleviated studies Oltjen (1967), Syrjälä (1972), and Camber- by increasing rumen pH, El-Shazly et al. lain et al. (1985) sucrose did not increase the (1961) proposed that when readily fermentable proportion of butyrate. The difference may carbohydrates are given, the competition for be related to the method by which the sugars essential nutrients is increased. Cellulolytic were given. microorganisms appear to be less competitive and unable to metabolise and reproduce at a rate fast enough to maintain themselves in the Conclusions rumen, and the number of cellulolytic bacteria falls. According to a review by VanGylswyk The present results indicate that production and Schwartz (1984), on the other hand, the of microbial protein from silage nitrogen can effect of readily fermentable carbohydrates on be increased by the continuous supply of fibre digestion is generally not due to a decline soluble carbohydrates. On silage diets the low in the number of cellulolytic bacteria. In the production of microbial N seems to be limited present study, the increased lag time of fibre by the energy supply per se rather than by the digestion with sugar infusions indicates that difference in the energy and nitrogen releases rumen microbes preferentially utilize soluble from silage. At the same time, a continuous carbohydrates before starting on the degrada- supply of readily fermentable carbohydrates tion of fibre (Mertens 1977). With sugar in- reduces fibre digestibility, although rumen pH fusions the fluid fraction contained approxi- is not affected. This would tend to reduce mately 0.60 of the total flow of microbial N silage DM intake and, hence utilization of from the rumen, while the corresponding silage. value for the control diet was 0.49. A similar effect was observed in the ratio of RNA content in the whole digesta to RNA content in Acknowledgements the solid phase. The ratio was 1.16 for control diet and 1.23 and 1.31 for low and high The author is grateful to Hannele Ala- Seppälä and M. Ranta for assistance sugar infusion. These observations indicate a technical during the experiment and to veterinarians shift in the ratio of free bacteria to attached Leena Oksanen and K. Skutnabb for surgical bacteria with sugar infusions which may in preparations of the experimental animals. part explain reduced fibre digestibility.

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SELOSTUS

Sakkarosi- ja xyioosi-infuusion vaikutus typen ja kuidun sulatukseen säilörehu-ohraruokinnalla

Pekka Huhtanen

Koe-eläiminä oli kaksi pötsi- jaohulsuolifistelöityä ay- fusoitaessa pötsiin sokeria 900 g/pv. Mikrobitypen vir- sonnia (elopaino 240 kg). Perusruokintana oli 12kg nur- taus ohutsuoleen lisääntyi lineaarisesti (P < 0.001) so- misäilörehua (227 g ka/kg ja 25.7 g N/kg ka), 1 kg litis- keritason lisääntyessä. Merkkiaineena käytettiin nukle- tettyä ohraa ja 100 g kivennäisseosta. Perusdieetin lisäk- iinihappojen puriiniemäksiä. Mikrobisynteesin tehokkuus si sonnit saivat jatkuvana infuusiona pötsiin 0, 450 ja (g N/kg pötsissä näennäisesti fermentoitunut OA) oli 24.0, 900 g/pv joko sakkaroosia tai xyloosia. 29.1 ja 30.0 sokeritasoilla 0, 450 ja 900 g/pv, vastaavas- Sokeriannosta lisättäessä orgaanisen aineen (OA) vir- ti. Pötsissä tuotetun mikrobitypen ja pötsinesteen taus ohutsuolen alkuosaan sekä sonnassa eritetyn OA:n NFlj-N-pitoisuuden välillä oli negatiivinen yhteys (r = määrä lisääntyivät lineaarisesti {P < 0.001); P < 0.01). —0.86; P < 0.001) sekä mikrobitypen ja nestevirtauk-

Pötsissä sulatetun OA:n osuus oli keskimäärin 0,694, ei- sen ohutsuolessa välillä positiivinen yhteys (r = 0.83; kä sokereiden tai sokeritasojen välillä ollut merkitseviä P <0.001). eroja. Neutraalidetergenttikuidun (NDF) ja happodetergent- Pötsinesteen NH,-N-pitoisuus ja isovaleriaanahapon tikuidun (ADF) virtaus ohutsuoleen (P < 0.01; P < osuus laskivat (P < 0.001) sokeriannoksen kasvaessa. 0.001) sekä eritys sonnassa (P < 0.05) lisääntyivät line- Sakkaroosi laski pötsinesteen NH,-N-pitoisuutla enem- aarisesti sokeriannosta lisättäessä. Sokereita infusoitaessa män (P < 0.05) kuin xyloosi. Sokeritaso ei vaikuttanut kuidun sulatus paksusuolessa lisääntyi, mutta tämä ei riit- etikka- propioni- ja voihapon osuuksiin pötsinesteen tänyt kompensoimaan huonontunutta kuidun sulatusta VFA:sta, mutta sakkaroosia infusoitaessa elikkahapon pötsissä ja siten NDF:n ja ADF:n kokonaissulavuus oli osuus oli pienempi (P < 0.05) ja voihapon osuus suu- huonompi (P < 0.05). Sokeriannoksen vaikutus nailon- rempi (P < 0.01) kuin xyloosia infusoitaessa. pussimenetelmällä määritettyyn typettömän orgaanisen Perusdieetillä ohutsuoleen virtaavan typen määrä oli aineen hävikkiin oli yhdenmukainen huonontuneen kui- 11.6 g/pv pienempi kuin rehussa saadun typen määrä, dun in vivo -sulavuuden kanssa, joskin vaikutus oli pie- mutta ylitti rehussa saadun typen määrän 21.6 g/pv in- nempi nailonpussimenetelmällä määritettäessä.

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