Coupling in vitro food digestion with in vitro epithelial absorption; recommendations for biocompatibility.

As food transits the gastrointestinal tract, food structures are disrupted and nutrients are absorbed across the gut barrier. In the past decade, great efforts have focused on the creation of a consensus gastrointestinal digestion protocol (i.e., INFOGEST method) to mimic digestion in the upper gut. However, to better determine the fate of food components, it is also critical to mimic food absorption in vitro. This is usually performed by treating polarized epithelial cells (i.e., differentiated Caco-2 monolayers) with food digesta. This food digesta contains digestive enzymes and bile salts, and if following the INFOGEST protocol, at concentrations that although physiologically relevant are harmful to cells. The lack of a harmonized protocol on how to prepare the food digesta samples for downstream Caco-2 studies creates challenges in comparing inter laboratory results. This article aims to critically review the current detoxification practices, highlight potential routes and their limitations, and recommend common approaches to ensure food digesta is biocompatible with Caco-2 monolayers. Our ultimate aim is to agree a harmonized consensus protocol or framework for in vitro studies focused on the absorption of food components across the intestinal barrier.

Fluid and particle retention in a small New World and a small Old World cervid, the southern pudu (Pudu puda) and Reeves's muntjac (Muntiacus reevesi).

Ruminants differ in the pattern how small particles and liquids pass through their gastrointestinal tract, and in particular their reticulorumen (RR). Based on that they may be classified into 'moose-type' and 'cattle-type' species (smaller and larger differences between particle and liquid passage, respectively). The ratio between the retention of particles and fluids is called the 'selectivity factor' (SF) and is a species-specific characteristic, studied in tragulids, giraffids and bovids, but not in many cervid species. Recently, it has been suggested that a high SF might also serve to wash digesta clean of external abrasives prior to regurgitation for rumination. In this study, we measured SF and passage kinetics (using a liquid marker and markers of different particle size, fed with the diet) in a capreoline deer, the southern pudu (Pudu puda, n = 5, 10.3 ± 2.9 kg, kept at two zoos) and a cervine deer, the Reeves's muntjac (Muntiacus reevesi, n = 6, 11.0 ± 1.7 kg, kept at a research facility). The relative daily dry matter intake (38 ± 3 g/kg0.75 for pudu and 76 ± 5 g/kg0.75 for muntjac) was higher, and the mean retention times (MRT) correspondingly shorter (e.g., MRT small particles in the total digestive tract 39 ± 8 h for pudu and 15 ± 2 h for muntjac), in the muntjac. The SF for small particles/liquid in the reticulorumen were, however, similar for both species, at 1.47 ± 0.21 for pudu and 1.66 ± 0.20 for muntjac, indicating a 'moose-type' physiology for both, irrespective of their different phylogenetic origin. To date, SF recorded in bovids attain distinctively higher values than the few reported for cervids. This situation reflects the degree of hypsodonty (tooth crown height) attained by these taxa, which is higher in bovids than in cervids. Together, constraints in hypsodonty as well as SF might limit cervids to more mesic habitats without distinct loads of external abrasives (such as dust or grit) on their food. In both species, some animals showed the typical ruminant pattern of a longer MRT for large than for small particle markers, but in some animals, this difference was not evident. This may be due to variable degrees of marker chewing during ingestion.

Individual differences in digesta retention and their relation to chewing in cattle-A pilot investigation.

While information on individual differences in digesta mean retention time (MRT) might be interesting when selecting phenotypes for digestive efficiency, MRT measurements are prohibitively labour-intensive for large-scale application. Therefore, more easily measured proxies of MRT might be helpful. We used the opportunity of an experiment applying saliva stimulant in cattle to investigate the effect of different individual chewing behaviour on fluid and particle MRT with a consistent diet. Four non-lactating cattle (670-850 kg body mass [BM]) were used in a 4 × 4 Latin square design, treated with the saliva stimulant pilocarpine in dosages of 0, 1, 2.5 and 5 mg/kg BM per day. The cattle were fed hay with dry matter intake (DMI) assigned according to their metabolic body weight. MRT in the whole gastrointestinal tract (GIT), the reticulorumen (RR) and the distal tract were measured using Co-EDTA, Cr-mordanted fibre and La-mordanted fibre as markers representing fluid, small particles (2 mm) and large particles (1 cm), respectively. The chewing behaviour was measured via noseband pressure sensor and expressed as chewing frequency (chews per time) and chewing intensity (chews per DMI), both for total chewing (ingestion plus rumination) and rumination chewing alone. The animals differed considerably in chewing behaviour and MRT measures. BM did not show a significant effect on chewing behaviour and MRT measures, though it tended to negatively correlated to total chewing intensity. Chewing intensity exerted a significant negative influence on MRT of fluid and particles in the RR, which was not the case for chewing frequency. Chewing frequency showed a significant relationship with MRT of large particles in the GIT. We suggest that chewing behaviour could influence MRT in two ways: (i) by affecting saliva production via the masticatory-salivary reflex and subsequently, the fluid inflow to the RR; (ii) by contributing to particle size reduction. Should the link between chewing behaviour and MRT be corroborated in larger studies, chewing measures, with their large interindividual variation, could emerge as an easy-to-measure proxy for MRT characteristics.

A high-fat and high-fructose diet lowers the cecal digesta's weight and short-chain fatty acid level of a Sprague-Dawley rat model.

BACKGROUND: This study aimed to analyze the effect of a high-fat and high-fructose diet (HFFD) on the digesta weight and shortchain fatty acid (SCFA) levels of cecal digesta in rats. METHODS: This study was an experimental study with a posttest-only control group design with male Sprague-Dawley strain rats as the samples. A total of 36 rats were divided into two groups with normal diet (N) and modified HFFD. The data of energy intake, nutrients and fiber, body weight, Lee index, abdominal circumference, digesta weight, and SCFA levels of cecal digesta were collected. RESULTS: The results showed an 11.94% increase in body weights of rats with HFFD. The total energy intake of the HFFD group was significantly higher than that of N (p = 0.000). The fiber intake and cecal digesta weight in group N were higher than that in the HFFD group (p = 0.00 and p = 0.02, respectively). The concentrations of acetate, butyrate, propionate, and total SCFA in the N group were significantly higher than in the HFFD (p = 0.041,,p = 0.004, p = 0.040, p = 0.013, respectively). A significant negative relationship was observed between the abdominal circumference and cecal digesta concentration (p = 0.029; r = -0.529) and between the Lee index and the SCFA concentration of cecal digesta (p = 0.036, r = -0.206).

Spatial and temporal organization of jejunal microbiota in goats during animal development process.

AIMS: This research aimed to investigate the temporal bacterial colonization relating to non-rumination, transition and rumination phases, together with the spatial organization of microbial community in the jejunal mucosa and digesta of goats. METHODS AND RESULTS: This study explored the colonization programme of the jejunal microbiota by employing 16S rRNA amplicon sequencing. The colonization pattern of jejunal bacterial community exhibited an age- and gut region-dependent progression during animal development process. Approximately 268 bacterial signatures contributed to the discrimination between gut regions, with Lactobacillus, Ruminococcus, Eubacterium and Clostridium_sensu_stricto were enriched in the jejunal digesta, and Bacteroides and unclassified bacteria were enriched in the jejunal mucosa. Intriguingly, a shift from Lactobacillus to Butyrivibrio, Eubacterium and Ruminococcus after d 20 was observed for jejunal digesta. In mucosa, Bifidobacterium, Corynebacterium, Faecalibacterium and Roseburia increased with age (P < 0·05) while Arcobacter, Bacteroides and Porphyromonas peaked at d 10. CONCLUSIONS: The jejunal bacterial community was settled after solid starter provision, which may mark the potential boundary of a timeframe for intervention in goats. The spatial heterogeneity highlighted the complicacy of ecological niches during manipulation of gut microbiota. SIGNIFICANCE AND IMPACT OF THE STUDY: The present study extended the understanding of microbial programming and niche specific in the jejunum among different life stages and the basal cognition of persistent enhancement of nutrient utilization and decline of enteric diseases in ruminants.

Comparison of the fermentation and bacterial community in the colon of Hu sheep fed a low-grain, non-pelleted, or pelleted high-grain diet.

Microbial fermentation in the hindgut is likely an important contributor to energy availability in ruminants, except for the rumen. This study aimed to investigate commensal bacteria in the colon influenced by diverse dietary niches. Fifteen male sheep were randomly allotted into three feeding groups: non-pelleted low-grain (CON, n = 5), non-pelleted high-grain (HG, n = 5), and pelleted high-grain (HP, n = 5) diets. The HG and HP groups had higher fermentation parameters than the CON group, especially acetate concentration (CON = 46.91; HG = 61.66; HP = 77.99). The HG diet altered the composition of commensal bacteria in the colon in comparison to the CON group, including the increase of genera related to acetate production (e.g., Acetitomaculum spp.), butyrate production (e.g., Coprococcus spp. and Subdoligranulum spp.), and starch degradation (e.g., Prevotella spp., Roseburia spp., and Oscillibacter spp.). The colon functional compendium had co-alteration with taxonomic changes that indicated non-pelleted HG diet caused a detrimental colonic niche. The HP diet specifically promoted the abundance of Ruminococcus, Olsenella, and Alloprevotella genera to achieve the highest acetate concentration and decreased the starch-degrader Roseburia spp. and Oscillibacter spp. in contrast to the HG group. Our results provide a systematic view of the microbial fermentation, community, and functional guilds in colonic digesta and mucosa in regard to using an HP diet to maintain colonic niche homeostasis under the adverse influence of the HG diet.Key Points• Non-pelleted and pelleted high-grain diets altered sheep colonic fermentation.• Non-pelleted and pelleted high-grain diets resulted in diverse microbial composition.• The pelleted method ameliorated microbial functions compared with the high-grain diet.

Nutrient digestibility and endogenous protein losses in the foregut and small intestine of weaned dairy calves fed calf starters with conventional or enzyme-treated soybean meal.

The aims of this experiment were (1) to compare the effects of a soybean meal with an enzymatic treatment (ESBM) to reduce the concentration of antinutritional factors versus a standard soybean meal (SBM) on foregut and small intestine digestion in weaned dairy calves and (2) to estimate the endogenous losses of crude protein (CP) in the small intestine. Our hypothesis was that a diet containing ESBM instead of SBM would improve ruminal and small intestine digestion and absorption of nutrients. A T-cannula was placed in the duodenum, and a second T-cannula was installed in the distal ileum of 12 Holstein calves at approximately 3 wk of age. Calves were weaned on d 42, and on d 50 they were assigned randomly to a quadruplicated 3 × 3 Latin square with 10-d periods. Digesta samples were collected on d 7 and 8 from the ileum and d 9 and 10 from the duodenum. The diets were fed for ad libitum intake and consisted of a calf starter (CS) of 20% CP with SBM as the main source of protein (CTRL), and an isonitrogenous CS with an ESBM instead of SBM (ENZT). A third diet with a low content of CP (10%) and no soy protein was fed to estimate endogenous N losses and digestibilities of test ingredients. Flows and digestibilities of nutrients were compared between CTRL and ENZT and their test ingredients (SBM vs. ESBM, respectively). Duodenal net flows of CP and total AA as well as ruminal microbial protein synthesis per kilogram of digested CP were greater, and flow of nonprotein N and CP true (corrected by endogenous and microbial flows) foregut digestibility were lower with ENZT than CTRL. The apparent small intestine digestibilities of CP and total AA were greater for ESBM than SBM, but there were no differences between the CTRL and ENZT diets. We observed no differences in digestibilities at the duodenum or ileum of starch or NDF, but true small intestine digestibilities of CP and all AA were greater with ENZT than CTRL. Total endogenous protein losses in the small intestine estimated from calves fed the low-CP with no soy protein diet were 37 ± 1.5 g of CP and 29 ± 1.4 g of AA/kg of DMI. These values may be considered the basal endogenous losses as they are similar to values obtained with the regression method, which estimates N losses when dietary N is null. Our results indicated that the inclusion of an ESBM improved the efficiency of ruminal microbial protein synthesis per digested kilogram of organic matter and CP, and increased CP and AA absorption in the small intestine despite a greater proportion of undigested dietary protein entering the duodenum.

Determination of colistin in luminal and parietal intestinal matrices of chicken by ultra-high-performance liquid chromatography-tandem mass spectrometry.

Justification for continued use of colistin in veterinary medicine, for example medicated water, relies on pharmacokinetic/pharmacodynamic (PK/PD) studies that require accurate measurement of colistin content in the digestive tract. A method for the detection and quantification of colistin in poultry intestinal material was developed and validated. Colistin is not absorbed after oral administration, and the biophase is the gastrointestinal tract. Extraction of colistin from the matrix was achieved using solid-phase extraction with a methanol:water (1:2; v/v) solution. Polymyxin B was used as an internal standard. Colistin A and colistin B, the main components of colistin, were separated, detected and measured using ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS). The method was validated for linearity/quadraticity between 1.1 (LOQ) and 56.7 mg/kg. Mean accuracy was between 82.7% and 107.7% with inter- and intra-day precision lower than 13.3% and 15% respectively. Freeze-thaw, long-term and bench storage were validated. Incurred samples following colistin treatment in poultry at the approved clinical dose of 75,000 IU/kg in drinking water and oral gavage were quantifiable and in line with expected intestinal transit times. The method is considered appropriately accurate and precise for the purposes of pharmacokinetic analysis in the gastrointestinal tract.

Effects of increasing dietary rye levels on physicochemical characteristics of digesta and its impact on stomach emptying as well as the formation of 'doughballs' in stomachs of young pigs.

Despite quite similar contents of starch and crude fibre of wheat and rye, the unique non-starch-polysaccharide fraction of rye (e.g. high levels of arabinoxylans and fructans) might have an impact on physicochemical properties of the digesta in pigs. Forty pigs (age: 46.8 ± 5.28 days; bodyweight: 16.1 ± 4.13 kg) were divided into four treatment groups. During four weeks, the pigs received diets consisting of wheat and/or rye, barley, soy, potato protein and a mineral supplement. The sum of wheat and rye was 69% in all diets, whereby the compound feed of each group was characterized by a different ratio (%) of wheat/rye (69/0; 46/23; 23/46; 0/69, respectively). In the stomach, 'doughballs' occurred more frequently with increasing dietary rye levels (9/10; 69% rye). With higher DM content and extract-viscosity of gastric digesta, the stomach emptying tended to be retarded in rye groups. Compared to the control group (69% wheat), maximum dietary rye levels (69%) resulted in significantly higher concentrations of lactic acid in digesta of the stomach and small intestine. With increasing lactic acid concentrations, the pH tended to be lower in small intestinal digesta. With an intensified formation of lactic acid, effects against Gram-negative bacteria, for example Salmonella, can be expected. Moreover, because of higher viscosity and the retarded stomach emptying, there could be advantages of including rye in compound feeds when a longer lasting satiety is intended, for example when feeding pregnant sows (regularly fed restrictively).

Cobalt (III)-EDTA dissociates and chromium (III)-EDTA is slightly more stable under in vitro reducing conditions comparable to those in the rumen.

Ideal digesta markers used for feeding studies are inert, unabsorbable, and move with the digesta they are intended to mark. Both chromium (III) and cobalt (III) salts of EDTA (CrEDTA and CoEDTA, respectively) are used as markers of liquid digesta in dairy cattle research. A small portion is absorbed and excreted in urine, but the markers are assumed to remain unreactive and as inert salts in the digesta and animal. The degree to which these colored salts remain bound in solution can be estimated through spectrophotometric measurement at their wavelength (λ) of peak absorbance. The objective of this in vitro study was to evaluate whether CrEDTA and CoEDTA dissociate under reducing conditions that could be experienced in the rumen. In a completely randomized design with 2 replicate analytical runs and samples in duplicate within run, approximately 26 mg/L Cr from CrEDTA or Co from CoEDTA was incubated in a 26-mL reaction volume containing 20 mL of Goering and Van Soest medium without tryptone, 3 mL of CoEDTA or CrEDTA solutions, or water (reagent blanks), and 3.0 mL of a combination of distilled water with 0, 0.25, 0.50, 0.75, or 1.00 mL of reducing solution (RedSol). After incubation for 0.5 h at 39°C, absorbance was read at λ = 535, 465, and 560 nm, the peak λ for EDTA salts of Co(III), Co(II), and Cr(III), respectively. Mean reagent blank values were subtracted from CoEDTA and CrEDTA data. The absorbance data at peak λ were analyzed by marker in models that included RedSol with analytical run as a random variable. Contrasts were used to detect linear through quartic effects of RedSol. Samples with RedSol had redox potentials of -250 to -328 mV, which are within the range of reported ruminal measures. As RedSol increased, CoEDTA showed a linear decline of 75% in ABS at 535 nm and a quadratic 4-fold increase followed by a 60% decline at 465 nm. These responses indicate a reduction of Co(III) to Co(II) and subsequent dissociation of Co(II)EDTA. The absorbance of CrEDTA at 560 nm showed a tendency for an 8% linear decrease as RedSol increased. Wavescans from λ = 330 to 700 nm showed CrEDTA retaining its characteristic 2-peak pattern as RedSol increased, whereas CoEDTA curves deformed entirely. We conclude that CoEDTA is not a stable, inert digesta marker under reducing conditions achievable in the rumen and is therefore unsuitable for use in studies with ruminants. Reexamination of the suitability of available liquid digesta markers is advised.

Short communication: A pilot study to describe duodenal and ileal flows of nutrients and to estimate small intestine endogenous protein losses in weaned calves.

The aims of this pilot study were (1) to evaluate the effect of an ileal and duodenal cannulation surgery on body weight and dry matter intake, (2) to estimate endogenous losses of crude protein (CP) and AA in the small intestine, and (3) to describe duodenal and ileal flows of nutrients in weaned dairy calves. Three Holstein male calves were fitted at 7 wk of life with a T-cannula at the terminal ileum and another cannula at the proximal duodenum. On wk 14 of life, calves were randomly assigned to a single 3 × 3 Latin square with 10-d periods. The 3 diets were fed ad libitum and consisted of a control calf starter (CS) with conventional soybean meal (SBM) as the main source of protein (CTRL), an isonitrogenous (20% CP) CS with an enzyme-treated SBM as the main source of protein (ENZT), and a CS with low content of CP (10%) and no soy protein (LOCP). Flows and digestibilities of nutrients were compared between the soy-based high-protein diets (HICP) and LOCP, and between CTRL and ENZT. Final data were only available from 2 calves per diet (n = 2) because cannulas from 1 calf became inoperative after the first collection period. Duodenal flows of CP, total AA, nonprotein nitrogen, microbial N, and fatty acids, as well as apparent duodenal digestibility of starch, were greater for HICP than for LOCP, indicating a greater foregut microbial activity and digestion. The apparent ileal digestibility (AID) of organic matter, CP, and total AA were greater for HICP than for LOCP. Duodenal net flow of CP was greater for ENZT than for CTRL, but flow of AA was not different. On the other hand, duodenal flow of microbial N was greater, and flows of nonprotein N and starch were lower for the ENZT diet, suggesting a more efficient microbial activity in the rumen. Even though CTRL had a greater AID when compared with ENZT, the AID of CP and AA were greater for enzyme-treated SBM than for SBM. Endogenous losses in the small intestine per kilogram of duodenal dry matter flow were 47 ± 15 and 37 ± 12 g/d, and the true ileal digestibilities for the HICP diets were 86 ± 0.1 and 87 ± 0.1% for CP and AA, respectively. An optimal supply of CP and the inclusion of an enzyme-treated SBM improved the efficiency of microbial digestion and increased AA absorption. Although further research with greater biological replication is needed, our results indicate that there is potential to improve digestion and absorption of proteins through dietary strategies in young weaned calves.

Effects of dietary inclusion of Zymomonas mobilis degraded cassava sifting on growth performance, apparent nutrient digestibility, ileal digesta viscosity, and economy of feed conversion of broiler chickens.

A study was carried out to evaluate the effect of dietary inclusion of Zymomonas mobilis degraded cassava sifting (ZDCS) on growth response, apparent nutrient digestibility, and ileal digesta viscosity of broiler chickens. Five diets containing undegraded and degraded cassava sifting were formulated to replace wheat offal at 0, 50, and 100% levels. Two hundred and forty (240) one-day-old Marshall broiler chickens were randomly allotted to the five dietary treatments in a completely randomized design (CRD); significant means were separated using Duncan's multiple range test at p < 0.05. The biodegradation of cassava sifting with Zymomonas mobilis significantly increased crude protein content by 44.59% while crude fiber and neutral detergent fiber significantly decreased by 23.08% and 6.38%, respectively. The results showed that birds fed 50% ZDCS had the best (p > 0.05) feed conversion ratio (FCR) at the starter phase. The replacement of wheat offal with 100% ZDCS improved (p < 0.05) the crude fiber digestibility at both starter and finisher phases. Also, the birds fed 100% ZDCS had the lowest (p < 0.05) value of ileal digesta viscosity. The birds fed 50% ZDCS had the highest (p < 0.05) values of gross revenue, gross profit, rate of return on investment, and economic efficiency while the least values for gross profit, rate of return on investment, and economic efficiency were obtained in 100% ZDCS. The study concluded that replacement of wheat offal with 50% ZDCS in the ration of broiler chickens improved FCR, crude fiber digestibility (CFD), and rate of return on investment and economic efficiency.

Whole digesta properties as influenced by feed processing explain variation in gastrointestinal transit times in pigs.

Physicochemical properties of diets are believed to play a major role in the regulation of digesta transit in the gastrointestinal tract. Starch, being the dominant nutrient in pig diets, strongly influences these properties. We studied transport of digesta solids and liquids through the upper gastrointestinal tract of ninety pigs in a 3 × 3 factorial arrangement. Dietary treatments varied in starch source (barley, maize and high-amylose maize) and form (isolated starch, ground cereal and extruded cereal). Mean retention times (MRT) of digesta solids ranged 129-225 min for the stomach and 86-124 min for the small intestine (SI). The MRT of solids consistently exceeded that of liquids in the stomach, but not in the SI. Solid digesta of pigs fed extruded cereals remained 29-75 min shorter in the stomach compared with pigs fed ground cereals (P < 0·001). Shear stress of whole digesta positively correlated with solid digesta MRT in the stomach (r 0·33, P < 0·001), but not in the SI. The saturation ratio (SR), the actual amount of water in stomach digesta as a fraction of the theoretical maximum held by the digesta matrix, explained more variation in digesta MRT than shear stress. The predictability of SR was hampered by the accumulation of large particles in the stomach. In addition, the water-holding capacity of gelatinised starch leads to a decreased SR of diets, but not of stomach digesta, which was caused by gastric hydrolysis of starch. Both of these phenomena hinder the predictability of gastric retention times based on feed properties.

Feed processing and structural components affect starch digestion dynamics in broiler chickens.

1. A 2 × 2 factorial design was used to test the hypothesis that impaired intestinal starch digestibility is attributable to rapid passage of digesta from the gizzard to the intestine, and that, compared to steam pelleting, increasing the availability of starch through extrusion cooking may alleviate the potential negative effect of rapid digesta flow on starch utilisation. 2. Thus, 7-d-old-broiler chickens were distributed to 48 cages and given a wheat-based (WB) pelleted diet containing either coarse oat hulls (OH-Pel) or fine cellulose (Cel-Pel) until d 19 to stimulate divergent development of the gizzard. Thereafter, both groups were further subdivided and challenged with a WB diet containing cellulose in either pelleted (Cel-Pel) or extruded (Cel-Ext) form on d 20 and 22. Either excreta or intestinal contents were collected at time intervals after feeding and analysed for marker and starch. 3. OH-Pel increased gizzard size and holding capacity. No excessively high starch levels (maximum 25 g/kg) were detected in the excreta. However, 8 h feed-deprived birds given Cel-Pel and challenged with Cel-Pel exhibited higher starch excretion and showed large individual variation during the first 135 min of collection. 4. Contrary to the OH-Pel group, more digesta and starch passed to the jejunum at 1 and 2 h and ileum at 2 and 3 h after feeding for birds given Cel-Pel, resulting in lower jejunal and ileal starch digestibility. 5. Increased starch gelatinisation through extrusion processing significantly improved starch digestibility regardless of gizzard function. However, at 1, 2 and 3 h after feeding, more digesta was retained in the foregut of birds given Cel-Ext. 6. The current data showed that starch degradation rate is associated with the flow of digesta which is linked to gizzard development, and that enzymatic hydrolysis of intact starch granules may be limited with more rapid feed passage through the gut.

Vegetable oil and carbohydrate-rich diets marginally affected intestine histomorphology, digestive enzymes activities, and gut microbiota of gilthead sea bream juveniles.

For an increased incorporation of plant ingredients in aquafeeds at the expense of fish meal (FM) and fish oil (FO), more knowledge is needed on the effects at the intestine level of dietary vegetable oils (VO) and carbohydrates (CH), and of possible interactions. For that purpose, in this study, the activities of digestive pancreatic enzymes (amylase, lipase, total alkaline proteases), gut microbiota, and histomorphology were assessed in gilthead sea bream (IBW 71.0 ± 1.5 g) fed four diets differing in lipid source (FO or a blend of VO) and carbohydrate content (0% or 20% gelatinized starch) for 81 days. No major changes in digestive enzyme activities were noticed in fish fed the experimental diets. Dietary VO, but not CH content, modified intestinal microbial profile, by increasing the similarity of bacterial communities. Especially when combined with CH, dietary VO promoted abnormal enterocyte architecture. Liver histology was also accessed, and an increased cytoplasmic vacuolization of hepatocytes was related with dietary CH inclusion, being only significantly different in fish fed FO-based diets. Overall, nutritional interactions between dietary lipid source and carbohydrate content were not observed on digestive enzyme activities and microbial profile. However, the intestine histological modifications observed in fish fed the VOCH+ diet suggest a negative interaction between dietary VO and CH. This requires a more in depth assessment in future studies as it can have negative consequences at a functional level.

Exploration of bimodal kinetics in marker digesta outflows using compartmental models.

A deterministic compartmental model is developed for examining the dynamics of digesta marker outflows in animals, focussing on models which will simulate bimodal kinetics with two peaks in the time course of marker outflow. First, to establish the background to the subsequent modelling of bimodal flow, we examine the compartmental scheme which gives rise mechanistically to the gamma function, and derive or describe various useful properties. This is illustrated by varying two key parameters of the gamma function, n (the number of compartments), and k (the rate constant for compartment emptying). Next, a more articulated compartmental scheme is constructed, and by progressive parameter changes, it is shown how bimodal faecal marker outflow can be achieved. Last, progressive simplification is applied to this scheme to arrive at what is (hopefully) the simplest compartmental scheme which can be used to simulate bimodal kinetics. This may be used mechanistically to describe the role of digesta flow in animals which exhibit such characteristics.

Comparative selective retention of particle size classes in the gastrointestinal tract of ponies and goats.

There is a discrepancy in the literature on potential digesta separation mechanisms in horses, with both a selective retention of fine and of large particles postulated in different publications. To assess the net effect of such mechanisms, we fed ponies on a hay-only diet a pulse dose of whole (unchopped) marked hay together with a solute marker, collected faeces on a regular basis, measured marker concentrations in whole faeces and in their large (2.0-16 mm), medium (0.5-1.0 mm) and small (0.063-0.25 mm) particle fraction, and calculated the corresponding mean retention times (MRTs). For comparison, the same experiment was performed in goats. In goats, as expected, MRTsolute (35 hr) was significantly shorter than MRTparticle (51 hr); only a very small fraction of particle marker was excreted as large particles (2%); and the MRT of these large particles was significantly shorter than that of small particles (with a relevant difference of 8.6 hr), indicating that those few large particles that escape the rumen do so mostly soon after ingestion. In ponies, MRTsolute (24 hr) did not differ from MRTparticle (24 hr); a higher fraction of particle marker was excreted as large particles (5%); and the MRT of these large particles was longer than that of small particles (but with a non-relevant difference of less than 1 hr). These results indicate that no relevant net separation of digesta phases occurs in horses and that selective particle retention mechanisms in the large intestine are unlikely to represent important characteristics of the horse's digestive physiology.

Effect of cereal straw and alfalfa hay diet on amino acid profile of gastrointestinal digesta in lactating dairy cows.

This study was conducted to evaluate the effects of replacing alfalfa hay with rice straw (RS) or corn stover (CS) on amino acid (AA) profiles of gastrointestinal digesta in lactating cows. Eighteen lactating dairy cows were randomly assigned to one of three groups (n = 6) and fed identical concentrate and corn silage with different forages on dry matter basis: (i) 23% alfalfa hay and 7% Chinese wild rye hay (AH); (ii) 30% CS; and (iii) 30% RS. After the 14-week feeding, a total of 18 cows were slaughtered to collect digesta from four representative organs, including rumen, omasum, duodenum and jejunum. The AA profiles of ruminal microbial fraction were similar among the treatments, except for greater Arg in cows fed RS than in cows fed AH or CS. Most of the analysed AA changed under different diets. Significant differences were found among the microbial fraction, rumen fluid and rumen digesta, with greater essential AA in digesta than in microbial fraction or rumen fluid and greater essential AA in microbial fraction than rumen fluid. Significant differences in individual AA profiles of digesta and relevant fluid were found across the four representative digestive tract parts, including rumen, omasum, duodenum and jejunum, showing much lower Leu proportion in CS and RS than in AH in duodenal fluid. In summary, ruminal microbes may prefer using essential AA, rather than non-essential AA. The AA profile of ruminal microbes was constant except for Arg. The AA composition of digesta across the four digestive tracts changed dramatically, which indicated differences in the ability and efficiency of AA absorption. The lower duodenum absorbable Leu proportion in cows fed CS or RS indicated the shortage of Leu in CS or RS diets, which might also restrict the balanced AA absorption.

Influence of feed provisioning prior to digesta sampling on precaecal amino acid digestibility in broiler chickens.

A regression approach was applied to determine the influence of feed provisioning prior to digesta sampling on precaecal (pc) amino acid (AA) digestibility in broiler chickens. Soybean meal was used as an example test ingredient. Five feed-provisioning protocols were investigated, four with restricted provision and one with ad libitum provision. When provision was restricted, feed was provided for 30 min after a withdrawal period of 12 h. Digesta were sampled 1, 2, 4 and 6 h after feeding commenced. A diet containing 300 g maize starch/kg was prepared. Half or all the maize starch was replaced with soybean meal in two other diets. Average pc digestibility of all determined AA in the soybean meal was 86% for the 4 and 6-h protocols and 66% and 60% for the 2 and 1-h protocols, respectively. Average pc AA digestibility of soybean meal was 76% for ad libitum feed provision. Feed provisioning also influenced the determined variance. Variance in digestibility ranked in magnitude 1 h > ad libitum > 2 h > 6 h > 4 h for all AA. Owing to the considerable influence of feed-provisioning protocols found in this study, comparisons of pc AA digestibility between studies applying different protocols prior to digesta sampling must be treated with caution. Digestibility experiments aimed at providing estimates for practical feed formulation should use feed-provisioning procedures similar to those used in practice.

Kinetics of solutes and particles of different size in the digestive tract of cattle of 0.5-10 years of age, and relationships with methane production.

The digestive physiology of cattle is characterised by comparatively long digesta mean retention times (MRTs), a particle sorting mechanism (difference in MRTs of large vs. small particles) and a distinct digesta washing (difference in MRTs between particles and fluids) in the reticulorumen (RR). How these processes mature during ontogeny, and how they link to other digestion characteristics and methane production, is largely unknown. We used a set of passage markers (Co-EDTA for fluids and hay particles of 2, 5 and 8 mm length mordanted with Cr, La and Ce, respectively) to measure MRTs in 12 heifers (0.5-2.1 years; hay only) and two groups of 15 lactating cows (2.4-10.0 years; forage-only vs. forage-concentrate diet). The MRTs differed between markers (Co < Cr < La < Ce) and were longer in heifers than cows, consistent with the lower feed intake in heifers. MRTs were mostly similar between cow groups and increased with age. Digesta washing was not affected by group, age, feed intake and number of chews per unit of feed. The degree of digesta washing was not related to CH4 measures. Particle sorting was more prominent in cows than heifers but did not differ between cow groups or change with age in cows. This could be the consequence of the abrupt increase in intake from heifers to cows at a time when gut capacity is not yet fully developed; particle sorting might then clear smaller particles from the RR sooner allowing a higher intake. Surprisingly, CH4 yield per ingested feed did not correlate with MRTs, and CH4 yield per unit of digested fibre decreased with increasing MRTs and with increasing fibre digestibility. As this pattern occurred in heifers and both cow groups, it appeared independent of age, indicating a mechanism that has not been described in the literature so far and requires further investigation.