Ammonia production , ammonia absorption , and urea recycling in ruminants . A review

Relevant research on ammonia production, ammonia absorption, and urea recycling in ruminants was reviewed. Ammonia production and utilization in the rumen and post-ruminal digestive tracts are described in detail. Absorption of ammonia into portal-drained viscera, ammonia detoxifi cation and urea synthesis in the liver, and urea degradation in the gastrointestinal tract are then discussed. The factors of affecting urea recycling and its pathways are also analysed. Suggested future research should focus on urea recycling dynamics and improvement of protein conversion effi ciency, and on the development of an integrated mechanistic model to describe the digestion and metabolism of nitrogen-containing compounds in ruminants fed practical diets.


INTRODUCTION
Ammonia and urea, in addition to amino acids (AA), peptides and microbial crude protein (MCP), etc., play an important role in nitrogen digestion and metabolism in ruminants.The feeding of non-protein nitrogen (NPN) supplements to ruminants is based on the knowledge that NH 3 is the major end-product of protein degradation in the rumen and on the belief, which appears to have been generally accepted, that most of the N utilized by rumen microbes comes from the NH 3 pool in the rumen (Nolan and Leng, 1972).Ruminants have been maintained on diets in which the only source of nitrogen was either NH 3 salts or urea, indicating that all of the AA that are essential for nonruminants can be synthesized by ruminal microorganisms.In ruminants, urea is an end product of N metabolism, but bacteria have the enzymatic capability to hydrolyze urea, thereby making urea-N available as NH 3 .The objective of this paper is to focus on certain aspects of urea recycling that have been reported and to relate these to their potential nutritional signifi cance in ruminants.

GENERAL OVERVIEW OF NH 3 PRODUCTION AND UREA RECYCLING
It is fi rst necessary to understand the complex and dynamic process of N digestion and metabolism in ruminants.Usually, ruminants are much less effi cient than nonruminants in utilizing high quality dietary proteins.A dominant feature of N digestion and metabolism is microbial conversion of some dietary protein to NH 3 in ruminants.Dietary intake protein (IP) is either degraded (DIP) in the rumen, with partial or total conversion to MCP, or passed from the rumen as undegradable intake protein (UIP).The DIP provides peptides, AA, and NH 3 to satisfy microbial requirements.In ruminants, NH 3 -N concentrations in the rumen usually surpass the practical requirements for microbial growth in the rumen; therefore, excess N is absorbed as NH 3 and converted to urea by the liver.Urea synthesized in the liver can diffuse into the rumen, small intestine, and hindgut, or be secreted in saliva for utilization by ruminal microbes.Urea is also excreted by the kidneys and present in other secretions.

AMMONIA PRODUCTION AND LOSS IN THE GASTROINTESTINAL TRACT
Ruminants absorb substantial amounts of their dietary N as NH 3 and, for many diets, more N is absorbed as NH 3 than as α-amino N (Reynolds, 1992).Ammonia generation in the gut results from two main processes: one is microbial degradation of nitrogenous compounds within the gut lumen, the other is microbial hydrolysis of urea passing across the gut wall from the blood and intestinal fl uids (Parker et al., 1995).The primary source of NH 3 within the rumen is dietary protein, except for ruminants consuming diets very low in protein.The UIP and indigestible intake protein (IIP) usually pass to the duodenum without affecting NH 3 production in the rumen.Degradable dietary NPN can be converted rapidly and quantitatively to NH 3 , dissolved nucleic acids in the rumen are also degraded extensively (Leng and Nolan, 1984) by rapid action of bacterial peptidases and deaminases to produce NH 3 .In addition, endogenous non-urea N and endogenous urea contribute to NH 3 sources of endogenous fermentable protein include sloughed mucosal cells and salivary proteins.Nolan (1975) indicated that 4.4 g of NH 3 -N per day were produced from these endogenous sources in sheep.Endo-genous urea can also serve as a signifi cant source of NH 3 in the rumen after either passing from blood plasma into the rumen or being swallowed in saliva (Leng and Nolan, 1984).Other routes of NH 3 production in the rumen include NH 3 derived from protozoa and fi xation of atmospheric N 2 , although the latter is apparently negligible (Li Pun and Satter, 1975).
Routes of NH 3 loss from the rumen pool include NH 3 -N incorporated into microbial cells, NH 3 outfl ow and NH 3 absorption.Al-Dehneh et al. (1997) noted that endogenous urea-N contributed 19.1 and 37.5% of the N in duodenal digesta and duodenal bacteria for lactating cows.Outfl ows of NH 3 depend on its concentration in the rumen and the fractional rate of fl uid outfl ow.Duodenal NH 3 fl ow measured in cows and sheep represented 2 and 9% of N intake, respectively (Firkins et al., 1987;Song and Kennedy, 1989).NH 3 absorption from the rumen is mainly a function of the ruminal concentration of NH 3 and is also the primary pathway of NH 3 loss from the rumen.Absorption does not appear to be by active transport but occurs via passive non-ionic diffusion down a concentration gradient (Parker et al., 1995).High concentrations of ruminal NH 3 increase the fl ux of NH 3 into the blood.Diffusion of NH 3 across the rumen wall has been demonstrated in vivo and in vitro (Siddons et al., 1985;Bödeker et al., 1990;Rémond et al., 1993).Under normal physiological conditions, most of the NH 3 in the gut lumen will be in the ionized form because its pH ranges from 2 to 6.
Usually, concentrations of NH 3 in hind-gut digesta are substantial.The NH 3 in the post-ruminal digestive tract includes NH 3 outfl ow from the reticulorumen, NH 3 from deaminated AA, and NH 3 from hydrolysed endogenous urea.Of these, much of the NH 3 comes from endogenous urea N in the post-ruminal digestive tract.Routes of NH 3 loss include incorporation of NH 3 into MCP, absorption of NH 3 , and elimination in faeces.Rémésy and Demigné (1989) found that NH 3 absorption from the caecum was increased in rats fed diets containing fermentable carbohydrates.Parker et al. (1995) concluded that this may in part be due to the increased entry of urea into the caecum and its hydrolysis by the caecal fl ora, it is also possible that the increased concentration of VFA in the caecal digesta had a more direct effect on NH 3 fl ux across the caecal wall.Meanwhile, it is also possible that bicarbonate has the ability to stimulate colonic NH 3 absorption in ruminants.Usually, bacteria in the digestive tract utilize NH 3 as their preferred source of N, and other forms of protein or AA are reduced to NH 3 before being used metabolically (Jackson, 1995).
UREA RECYCLING Urea recycling is signifi cantly related to NH 3 production and absorption in the gastrointestinal tract (GIT) of ruminants.All NH 3 absorbed from the rumen epithelium, small intestinal mucosa, and large intestinal mucosa travels via the portal vein to the liver; body tissue NH 3 also enters the liver.Liver metabolism has a central role in the integration of body N metabolism.Ammonia in the liver is detoxifi ed by conversion to urea, urea can then be recycled directly into the rumen, small intestine, or large intestine; it can enter the rumen in saliva, be excreted by the kidney, or be secreted in milk or sweat (Alio et al., 2000).
Although portal absorption rates provide an overall measure of NH 3 fl ux into the blood, a number of different techniques have been used to study the relative contribution of different sections of the digestive tract to total NH 3 absorption by PDV (Parker et al., 1995).Siddons et al. (1985) provided a dynamic model of NH 3 -N transfer across different sections of the digestive tract.Seal and Reynolds (1993) probed the relationship between NH 3 fl ux in portal blood and dietary N intake; they found that portal NH 3 fl ux can represent as much as 0.65 of N intake and in many circumstances can exceed net α-NH 2 -N absorption into portal blood.The NH 3 fl ux from different sections of the gastrointestinal tract can be measured using chronically-catheterized animals.Body tissue NH 3 fl ux into the liver can then be deduced, provided that the overall rate of urea synthesis from NH 3 is known.
HEPATIC DETOXIFICATION OF NH 3 , UREA SYNTHESIS AND REMOVAL Ammonia is extremely toxic in non-hepatic tissues, causing changes in cerebral metabolism that can result in tetany and death when circulating concentrations exceed 0.7 mM (Symonds et al., 1981).Under normal physiological and nutritional conditions, NH 3 absorbed into the portal vein is effi ciently extracted by the liver and detoxifi ed by conversion to urea or glutamine.Over a wide range of portal NH 3 concentrations and on a variety of diets, the liver is able to extract 70 to 95% of portal NH 3 .As a result, hepatic NH 3 removal is on average slightly higher (4%) than portal absorption.Thus, arterial NH 3 concentrations remain relatively constant even when portal NH 3 absorption varies threefold (Parker et al., 1995).This ensures that any NH 3 which escapes conversion to urea in periportal hepatocytes is converted to glutamine in perivenous hepatocytes.Amide-N of glutamine is then removed and metabolized to urea by periportal hepatocytes during subsequent passages through the liver, and may also provide a mechanism to avoid a decrease in extracellular pH (Haussinger et al., 1992).Havassy et al. (1982) and Kowalczyk et al. (1982) found that urea nitrogen in the rumen was fi xed transiently into plasma protein.Enrichment of 15 N of the bacterial matter and plasma protein exceeded that of individual amino acid indicating that urea nitrogen was utilized to a large extent for the synthesis of nitrogen compounds other than amino acids.Maltby et al. (1991Maltby et al. ( , 1993b) ) reported that when urea was added to ruminant diets there was increased hepatic NH 3 uptake but glutamine uptake was either unchanged or slightly increased; however, net hepatic output of glutamate was decreased.The conversion of NH 3 to glutamine and glutamate TAN Z., MURPHY M. R.
is not a major detoxifi cation pathway under normal feeding conditions.Lobley et al. (1995) reported that 93.5 and 6% of portal 15 NH 4 Cl is converted to 15 N-urea and 15 N-glutamine, respectively, when portal vein NH 3 concentrations were increased to 0.5 mM by intramesenteric vein infusion in sheep.The capacity of ruminant liver to remove NH 3 is apparently 1.2 to 1.5 µmol/min per gram (Symonds et al., 1981) and the potential contribution of extracted NH 3 -N to hepatic urea-N formation ranges from 27 to 110%.Reasons for variation in the contribution of NH 3 to hepatic urea production are not clear, but Parker et al. (1995) considered the large range not to be an artifact and noted that animal factors and intake appeared not to be implicated.The major route of urea entry from blood to the rumen is via saliva.

UREA DEGRADATION IN THE RETICULORUMEN AND POST-RUMINAL DIGESTIVE TRACTS
Substantial amounts of recycled urea-N can be used by bacteria in the lumen of the gut for metabolic needs or reabsorbed as N in the forms of AA, nucleic acids, or NH 3 (Kowalczyk et al., 1975a;Nolan and Stachiw, 1979;Huntington, 1989;Reynolds, 1992).This provides a mechanism for salvage of urea-N by conversion into bacterial matter that can then be digested, yielding AA for use by the host (Sarraseca et al., 1998).Jackson (1995) concluded that urea-N retained in the body might, in principle, be converted into AA-N in one of four ways: absorbed as NH 3 and fi xed in the liver through amination to form non-essential AA, e.g., as glutamate or glycine-serine; through wider transamination with the C skeleton of a transaminating non-essential AA, e.g., alanine and aspartate; through wider transamination with the C skeleton of a transaminating essential AA; by bacterial synthesis of an essential or non-essential AA.
The amount of urea-N transferred into the rumen is determined by the rate of salivary secretion and by the plasma urea concentration.Nolan and MacRae (1976) reported that 5.3 g of blood urea-N/d entered the digestive tract of sheep; 20% of this urea was degraded in the rumen, 25% in the caecum, and the remainder was apparently degraded elsewhere.There was evidence of urea degradation in the large intestine posterior to the caecum, and it was suggested that urea degradation and absorption of the synthesized NH 3 might also occur in the ileum.Kowalczyk et al. (1975a,b) stated that only a small amount of blood urea nitrogen was utilized for microbial synthesis in the rumen, and the greatest part of postruminal endogenous nitrogen was reabsorbed during passage of digesta through the intestine (Sandek et al., 2002).Norton et al. (1978) noted that an average of 81% of the urea synthesized in the body was transferred to the digestive tract and degraded to NH 3 and carbon dioxide.Endogenous urea degraded in the rumen accounted for 7 to 13% of the total quantity degraded in the digestive tract, and the rate of urea transfer was not related to the rate of urea synthesis in the body.The lower digestive tract was the major site of urea degradation in sheep given low protein diets, and the rate of urea transfer to this part of the digestive tract was linearly related to the rate of urea synthesis in the body.Koenig et al. (2000) and Newbold et al. (2000) reported that urea-N contributed 20% of rumen NH 3 fl ux in sheep offered either a forage-concentrate ration or pelleted dried grass, respectively.It was concluded that urea transferred from the blood to the reticulo-rumen and the hind gut is degradable, making NH 3 -N available for use by microorganisms or for reabsorption and utilization by the body.

FACTORS AFFECTING UREA RECYCLING
All factors that infl uence the production, absorption, and transfer of NH 3 and urea will affect urea recycling in ruminants.Kennedy and Milligan (1980) reported that urea transfer to the rumen was inversely related to the rumen NH 3 concentration, and suggested that the NH 3 concentration was a factor regulating urea entry into the rumen.There was a marked reduction of urea transfer to the rumen when the ruminal NH 3 concentration was elevated by continuous NH 3 infusion into it.Rémond et al. (1993) concluded that NH 3 absorption seems to be mainly infl uenced by the NH 3 concentration in the rumen fl uid, and by the rate of VFA absorption.Net NH 3 fl ux across the rumen wall is linearly related to both free NH 3 and to total NH 3 concentrations.Additionally, ruminal VFA may stimulate the uptake of NH 3 .Bödeker et al. (1992) found that NH 3 absorption was stimulated by the presence of VFA in the mucosal buffer solution, either individually or as a mixture of acetate, propionate, and butyrate.Similar responses to additional butyrate on transfer of NH 3 into the ruminal vein of sheep have also been reported (Rémond et al., 1993).
Tracer studies have indicated that a supplemental energy source such as grain, starch, or dried beet pulp, signifi cantly increased endogenous urea degradation in the gastrointestinal tract.It is possible that the rumen was the site of increased degradation because Kennedy and Milligan (1980) reported that dietary sucrose greatly enhanced the rate of transfer of urea to the rumen.Huntington and Reynolds (1986) pointed out that the effects of dietary energy density or fermentability of a substrate in the rumen on the rate and site of endogenous urea transfer to the gut were obvious.
On the other hand, urea transfer from blood to the GIT might not be controlled by the urea concentration in plasma alone.In sheep and cattle, the upper limits of the blood urea concentration above which urea transfer was no longer linearly related to plasma urea concentrations were 6.0 mM and 4.0 mM, respectively.Elevation of plasma urea above these concentrations did not further increase rumen NH 3 .Norton et al. (1978) found that transfer of urea into the post-ruminal tract is correlated with both plasma urea concentration and its production rate.
Feed intake is an important factor that infl uences the return of urea to the GIT.Bunting et al. (1989) reported that net incorporation of blood urea-N into bacterial protein is inversely related to the protein intake of calves.Sarraseca et al. (1998) found that urea-N production in sheep increased with intake and exceeded digestible N at all intakes.Urea-N entering the digestive tract that was returned to the ornithine cycle remained constant across intakes but the absolute amount increased with N intake.Urea removed by the PDV, unaffected by intake, represented 32, 33, and 21% of the digested N. Meanwhile the reabsorption of endogenous nitrogen was signifi cantly infl uenced by the dietary crude fi bre level for growing sheep (Sandek et al., 2002).Leng and Nolan (1984) reported that NH 3 concentrations in rumen fl uid were positively correlated with the number of ciliate protozoa, and NH 3 concentrations in the rumen fl uid of defaunated animals were lower than in those with protozoa.The absorption of NH 3 is also likely to be lower in defaunated animals.Additionally, Allan and Miller (1976) found that, at equal rates of urea production, lambs tended to maintain higher plasma urea concentrations and greater rates of urea degradation in the GIT than did wethers; urea degradation was related to plasma urea concentrations in lambs but not in wethers.
Lastly, there are other factors that can also infl uence urea recycling for ruminants, such as physiological conditions, osmolality in the GIT, gastrointestinal hormones, etc. (Kennedy and Milligan, 1980).Bödeker et al. (1991) found that HCO 3 -favoured NH 3 absorption across the ruminal epithelium and Rémond et al. (1993) noted that CO 2 insuffl ation resulted in a 16% increase in net transfer of NH 3 across the ruminal wall.Although blood fl ow to the sheep rumen was increased, increased osmolality after NaCl injection slightly decreased NH 3 absorption.

RESEARCH APPROACHES
Two approaches are often used to study the recycling of nitrogen-containing compounds.One involves use of labelled isotope tracers and the other employs chronic catheterization of venous and arterial vessels.

Single or continuous isotope tracer technique
First, it is necessary to assume that the animal is in a steady state; i.e. pool sizes remain constant and the rates of infl ow and outfl ow are equal.Isotope tracer techniques generally include both single infusion and continuous infusion methods.Most of the metabolic studies made with 15 N have used single infusion rather than continuous infusion methods.
Analysis of isotope ratio with time yield curves for various primary compartments.The change in isotope ratio (Y t ) in a primary pool with time after a single injection of an isotope tracer is given by a multi-exponential curve of the form: where t = time, A i = 15 N-enrichment in the corresponding pool at the zero-time intercept of the i th compartment, m i = the fractional rate constant for the i th compartment, n = the number of exponential compartment, i = the exponential compartment number, and Y t = 15 N-enrichment in the corresponding pool at time t.
Parameter estimates from the fi tted equations are then used to calculate ruminal NH 3 -N pool size, total entry rate or fl ux, irreversible loss, and recycling.The relevant equations are as follows: where Q = pool size, D = dose of 15 N, F = total fl ux rate, a i is the fractional zerotime intercept of component A i , L = irreversible loss rate, R = recycling rate.
The quantity of urea synthesized in the body, degraded to NH 3 in the digestive tract, and subsequently resynthesized into urea by the body can be estimated from the difference between the rates of irreversible loss of urea-C and urea-N from plasma, as estimated by using simultaneous injection of 14 C-urea and 15 N-urea.This is possible because 14 C from hydrolysed urea enters a very large bicarbonate pool with a rapid turnover; therefore, the return of 14 C into newly synthesized urea is negligible (Koenig et al., 2000).As for the continuous injection method, the irreversible loss rate of NH 3 from ruminal fl uid is calculated by comparing enrichment of NH 3 at "plateau" enrichment with the rate of infusion of 15 N ammonium sulphate.The proportion of urea in plasma, or the bacteria-N in the ruminal fl uid derived from this, is calculated as the ratio of the "plateau" enrichments of urea-N or bacteria-N to NH 3 -N.
Recently, a different isotope tracer technique has been developed (Sarraseca et al., 1998).This method involves infusion of ( 15 N 15 N)-urea, followed by isotope analysis of three species ( 15 N 15 N), ( 14 N 15 N) and ( 14 N 14 N).The technique is based on the assumption that when urea enters the GIT as a ( 15 N 15 N) molecule and then undergoes hydrolysis via bacterial urease action, this will yield two molecules of 15 NH 3 .If these 15 NH 3 molecules are then reabsorbed and extracted by the liver then they may com- bine with 14 N atoms (from aspartate) within the hepatic ornithine cycle to yield two ( 15 N 14 N)-urea molecules.The chances of ( 15 N 15 N)-urea returning to the system after entry to the gut, whether directly or indirectly by combination of two 15 N-containing molecules within the ornithine cycle, are considered negligible.

Arterial-venous difference techniques
This technique requires precise surgical interventions, liver and GIT metabolism can be separated and the latter can be further separated via careful catheterization; accurate determination of blood fl ows is required.Chronic catheters are usually inserted into the hepatic vein, the portal vein, a mesenteric vein, the ruminal vein and a jugular artery.Rates of NH 3 absorption and urea synthesis can be estimated but data on the fate of urea-N are not (Table 1).According to Table 1, the following relationships can be obtained between N intake (x 1 in g per day) or portal NH 3 absorption (x 2 in mmol per min) and hepatic urea synthesis (y in mmol per min): Cattle: y = -116.8+ 2.94x 1 , r 2 = 0.77, n = 20 y = 13.0 + 1.53x 2 , r 2 = 0.77, n = 20 Cow: y = 1171.8− 1.08x 1 , r 2 = 0.26, n = 4 y = -143.0+ 1.72x 2 , r 2 = 0.88, n = 4 Sheep: y = 29.0+ 0.60x 1 , r 2 = 0.06, n = 10 y = 11.1 + 1.23x 2 , r 2 = 0.10, n = 10 It was concluded that signifi cant relationships exist between dietary N intake or portal NH 3 absorption and hepatic urea synthesis in cattle.In sheep, much weaker relationships are noted between N intake or portal NH 3 absorption and hepatic urea synthesis.

MANIPULATING PATHWAYS OF UREA RECYCLING
In ruminants, especially those in developing countries, protein nutrition becomes more important because protein resources are limited.In developed countries N in urine and faeces has sometimes become a serious environmental burden because of extensive use of high protein diets.Improvement of the utilization effi ciency of dietary protein and reduction of N waste through rational formulation of diets has become very important.A considerable portion of N excreted in the faeces is endogenous material.Urinary N originates from inevitable losses related to maintenance, losses associated with the deposition of AA into skeletal muscle tissue, losses resulting from any imbalance between energy and protein supplied by the diet, and excretion of purine derivatives, degradation products of microbial  High grain diets-different RDP Krehbiel and Ferrell (1999) 9.5% CP nucleic acids absorbed from the small intestine (Van Bruchem et al., 1997).Excretion of endogenous N is considerably higher than ileal N fl ow or the quantity eliminated in faeces.These fractions only constitute part of the total quantity of endogenous protein produced because much endogenous protein is reabsorbed.Of these routes, urea recycling is one of the most important components of endogenous protein reabsorption.The transport of urea N into the GIT is a normal process of great signifi cance in the physiology of all mammals.A large amount of N passes through the urea pool daily and this provides a potential target for manipulation to improve its conversion to animal products.There are two main directions: either reduce the amount of dietary N converted to urea by reducing NH 3 absorption and AA catabolism, or improve the conversion of urea-N, produced in the liver and returned to the digestive tract, into microbial protein.Although many complicated relationships exist between N intake and hepatic urea-N (Table 1), Bunting et al. (1989) noted that net incorporation of blood urea-N into bacterial protein is inversely related to N intake; this implies that hepatic NH 3 production may be reduced by manipulating dietary N sources.For example, a less degradable source of dietary protein may provide a greater proportion of absorbable AA entering small intestines; this can reduce urea synthesis.Krehbiel and Ferrell (1999) noted that for cattle consuming high-grain diets, an optimal amount of DIP in the diets enhanced fermentation in the rumen, increased AA fl ow to the duodenum, and increased net portal appearance of AA, without infl uencing energy use by the PDV.When the dietary DIP requirement has been met, additional DIP will not be benefi cial.For example Bohnert et al. (1999) demonstrated that N retention and effi ciency were improved by increasing UIP from 40 to 60% of total CP when lambs were fed low energy diets.
On the other hand, supplementing diets with fermentable energy sources can enhance utilization of N for bacterial synthesis.Huntington (1989) reported that increased intake of readily fermentable carbohydrate increased the rate of endogenous urea transfer through the rumen wall, decreased salivary transfer of urea to the rumen, and decreased urea transfer to post-ruminal tissues.Thus, MCP production in the rumen can be increased.Obitsu et al. (2000) reported that abomasal infusion of glucose reduced urea production and urinary N excretion.This illustrates that increased glucose absorption from the small intestine may contribute to increased fl ow of AA to peripheral tissues and to reduced wastage as excretion of urinary N. As described above, it is necessary to maintain a suitable ratio of available energy and N to improve the utilization of recycled urea.
It is well known that protozoa in the rumen increase the degradation of dietary protein, producing a rapid release of NH 3 .They also ingest and digest bacteria, recycling more N. Jouany (1996) concluded that defaunation may improve N utilization in ruminants by increasing AA supply to the intestines, by reducing TAN Z., MURPHY M. R. urea synthesis, and by reducing urinary N excretion and bacterial N turnover.In addition, Koenig et al. (2000) demonstrated that defaunation improved the intraruminal metabolism of N by increasing both the ruminal concentration of bacteria and the fl ow of bacterial N to the intestine.
It is also possible to manipulate the utilization effi ciency of recycled urea by adjusting diet structure (i.e. the forage to concentrate ratio) and intake.Huntington et al. (1996) reported that, when fed diets with 20% or less concentrate, steers recycled 90% of hepatic urea production; the percentage of recycled urea decreased to 64 with 63% of the diet as concentrates, and to 51% when diets with 90% concentrate were fed.Increased urea recycling to the GIT may improve the overall effi ciency of N utilization for maintenance and production.Under conditions of low or zero N intakes, urea-N production exceeds N intake in ruminants as body protein is mobilized when animals are in negative N balance.

FUTURE DIRECTIONS
Much data on NH 3 production, NH 3 absorption, and urea recycling in ruminants has been accumulated (Table 1); however, it is still diffi cult to utilize these data in feeding practice because few experiments have been conducted using practical diets.In the future, we propose that nutritional manipulations of N utilization must be tested on, and applied to, practical diets while the database on this topic continues to grow, i.e. while the optimal nitrogen intake, the ratio of dietary nitrogen and readily fermentable carbohydrate, are determined by the slow release technique for NPN in the rumen and the bypass technique for crude protein.After obtaining these practical data, these techniques or nutritional parameters will be organically conformed and applied in the practical diets of various animals.Further, considering the complexity of urea recycling, it is necessary to integrate the effects of the many dietary factors involved and to study their dynamics.This will involve the development and refi nement of mechanistic metabolic models.
urea-N output (H-Urea) (mmol/h) measured using the AV difference technique in ruminants fed a range of diets Species