Evaluation of titanium dioxide and chromic oxide as digestibility markers in ponies fed alfalfa hay in relation to marker dosing frequency.

In equines, Cr2O3 is widely accepted as an indigestible marker, but there are health concerns regarding the carcinogenic properties of Cr2O3. Recently, TiO2 has been suggested to be an alternative digestibility marker in equines. However, a comparison between Cr2O3 and TiO2 has not been made in equines. Six Welsh pony geldings (initial BW: 254±3 kg; 7 years of age) fed chopped alfalfa hay were used to evaluate the use of TiO2 (Ti) and Cr2O3 (Cr) as markers for calculating apparent digestibility and to investigate the effect of frequency of marker administration on the measurement of digestibility values. Diets contained 4.65 kg dry matter (DM) chopped alfalfa hay supplemented with minerals, vitamins, TiO2 (3.3 g Ti/day) and Cr2O3 (3.2 g Cr/day). Ponies were dosed with either 3.3 g Ti and 3.2 g Cr once daily (DF1) or with 1.65 g Ti and 1.60 g Cr twice daily (DF2). After adaptation to the diets and procedures for 14 days, voluntary voided faeces were collected quantitatively over 7 days and analysed for moisture, ash, Ti and Cr. Apparent total tract DM digestibility (DMD) and organic matter digestibility (OMD) were calculated using the total faecal collection (TFC) and marker method (Ti and Cr). The overall mean cumulative faecal recovery of Cr and Ti (as % of intake) were 102.0% and 96.6%, respectively. Mean daily faecal recoveries of Cr as well as of Ti were not different (P=0.323; P=0.808, respectively) between treatments. Overall daily faecal recovery of Cr differed (P=0.019) from 100% when the marker was dosed once daily, whereas overall daily faecal recovery was similar to 100% for both administration frequencies when Ti was used as a marker. For both markers, the coefficient of variation of the mean faecal marker recovery between horses was lower when the markers were administrated twice per day. Across treatments, cumulative DMD and OMD estimated with Ti were similar (P=0.345; P=0.418, respectively) compared with those values determined by TFC method. When Cr was used, the calculated cumulative DMD tended (P=0.097) to be greater compared with those estimated with TFC, and cumulative OMD values were overestimated (P=0.013). Orally supplemented Ti recovery in the faeces of ponies fed chopped alfalfa hay with Ti administered once or twice daily was close to 100%, making it the preferred marker for digestibility trials in equines.

Effect of exercise on apparent total tract digestibility of nutrients and faecal recovery of ADL and TiO2 in ponies.

Exercise and physical training are known to affect gastrointestinal function and digestibility in horses and can lead to inaccurate estimates of nutrient and energy digestibility when markers are used. The effect of exercise on apparent nutrient digestibility and faecal recoveries of ADL and TiO2 was studied in six Welsh pony geldings subjected to either a low- (LI) or high-intensity (HI) exercise regime according to a cross-over design. Ponies performing LI exercise were walked once per day for 45 min in a horse walker (5 km/h) for 47 consecutive days. Ponies submitted to HI exercise were gradually trained for the same 47 days according a standardized protocol. Throughout the experiment, the ponies received a fixed level of feed and the daily rations consisted of 4.7 kg DM of grass hay and 0.95 kg DM of concentrate. The diet was supplemented with minerals, vitamins and TiO2 (3.0 g Ti/day). Total tract digestibility of DM, organic matter (OM), CP, crude fat, NDF, ADF, starch, sugar and energy was determined with the total faeces collection (TFC) method. In addition, DM and OM digestibility was estimated using internal ADL and the externally supplemented Ti as markers. Urine was collected on the final 2 days of each experimental period. Exercise did not affect apparent digestibility of CP, crude fat, starch and sugar. Digestibility of DM (DMD), OM (OMD), ADF and NDF tended to be lower and DE was decreased when ponies received the HI exercise regime. For all treatments combined, mean faecal recoveries of ADL and Ti were 87.8±1.7% and 99.3±1.7%, respectively. Ti was not detected in the urine, indicating that intestinal integrity was maintained with exercise. Dry matter digestibility estimated with the TFC, ADL and Ti for ponies subjected to LI exercise were 66.3%, 60.3% and 64.8%, respectively, while DMD for HI ponies were 64.2%, 60.3% and 65.2%, respectively. In conclusion, physical exercise has an influence on the GE digestibility of the feed in ponies provided with equivalent levels of feed intake. In addition, the two markers used for estimating apparent DMD and OMD indicate that externally supplemented Ti is a suitable marker to determine digestibility of nutrients in horses performing exercise unlike dietary ADL.

Evaluation of methodological aspects of digestibility measurements in ponies fed different haylage to concentrate ratios.

Methodological aspects of digestibility measurements were studied in four Welsh pony geldings consuming haylage-based diets with increasing proportions of a pelleted concentrate according to a 4×4 Latin square design experiment. Ponies were fed four experimental, iso-energetic (net energy (NE) basis) diets (i.e. 22 MJ NE/day) with increasing proportions of a pelleted concentrate (C) in relation to haylage (H). The absolute amounts of diet dry matter fed per day were 4.48 kg of H (100H), 3.36 and 0.73 kg of H and C (75H25C), 2.24 and 1.45 kg of H and C (50H50C) and 1.12 and 2.17 kg of H and C (25H75C). Diets were supplemented with minerals, vitamins and TiO2 (3.7 g Ti/day). Voluntary voided faeces were quantitatively collected daily during 10 consecutive days and analysed for moisture, ash, ADL, acid-insoluble ash (AIA) and Ti. A minimum faeces collection period of 6 consecutive days, along with a 14-day period to adapt the animals to the diets and become accustomed to the collection procedure, is recommended to obtain accurate estimations on dry matter digestibility and organic matter digestibility (OMD) in equids fed haylage-based diets supplemented with concentrate. In addition, the recovery of AIA, ADL and Ti was determined and evaluated. Mean faecal recovery over 10 consecutive days across diets for AIA, ADL and Ti was 124.9% (SEM 2.9), 108.7% (SEM 2.0) and 97.5% (SEM 0.9), respectively. Cumulative faecal recovery of AIA significantly differed between treatments, indicating that AIA is inadequate to estimate the OMD in equines. In addition, evaluation of the CV of mean cumulative faecal recoveries obtained by AIA, ADL and Ti showed greater variations in faecal excretion of AIA (9.1) and ADL (7.4) than Ti (3.7). The accuracy of prediction of OMD was higher with the use of Ti than ADL. The use of Ti is preferred as a marker in digestibility trials in equines fed haylage-based diets supplemented with increasing amounts of pelleted concentrate.

The nutritive value of condensed wheat distillers solubles for cattle.

The chemical composition and the energy and protein value of five batches of condensed distillers solubles (CDS) originating from wheat were determined. The net energy for lactation (NEL) was derived from digestion coefficients obtained with sheep. The true protein digested in the small intestine (DVE) and the rumen degradable protein balance (OEB) were based on the rumen degradation rate (kd D ), the rumen undegradable fraction (U) and intestinal digestibility of undegraded protein (%DVBE) predicted by regression equations derived from a data set of 28 protein feeds with kd D , U and %DVBE determined in situ. The CDS is a by-product with a high, but very variable CP content (238 to 495 g/kg DM). The CP contained on average 81% amino acids, with glutamine as main component (on average 21.8% of CP) and a relatively good lysine proportion (3.0%). Further, CDS contains quite a lot of crude fat (mean±SD: 71±14 g/kg DM), glycerol (95±52 g/kg DM) and sugars (123±24 g/kg DM) resulting in a high organic matter digestibility (88.6±3.0%) and high NEL content (8.3±0.4 MJ/kg DM). The protein value showed a large variation, with DVE ranging from 122 to 244 g/kg DM and OEB from 50 to 204 g/kg DM. Wheat CDS is a rich source of minerals and trace elements with exception of calcium.

Comparison of fractionation methods for nitrogen and starch in maize and grass silages.

In in situ nylon bag technique, many feed evaluation systems use a washing machine method (WMM) to determine the washout (W) fraction and to wash the rumen incubated nylon bags. As this method has some disadvantages, an alternate modified method (MM) was recently introduced. The aim of this study was to determine and compare the W and non-washout (D+U) fractions of nitrogen (N) and/or starch of maize and grass silages, using the WMM and the MM. Ninety-nine maize silage and 99 grass silage samples were selected with a broad range in chemical composition. The results showed a large range in the W, soluble (S) and D+U fractions of N of maize and grass silages and the W, insoluble washout (W-S) and D+U fractions of starch of maize silages, determined by both methods, due to variation in their chemical composition. The values for N fractions of maize and grass silages obtained with both methods were found different (p < 0.001). Large differences (p < 0.001) were found in the D+U fraction of starch of maize silages which might be due to different methodological approaches, such as different rinsing procedures (washing vs. shaking), duration of rinsing (40 min vs. 60 min) and different solvents (water vs. buffer solution). The large differences (p < 0.001) in the W-S and D+U fractions of starch determined with both methods can led to different predicted values for the effective rumen starch degradability. In conclusion, the MM with one recommended shaking procedure, performed under identical and controlled experimental conditions, can give more reliable results compared to the WMM, using different washing programs and procedures.

Evaluation of methodological aspects of digestibility measurements in ponies fed different grass hays.

Methodological aspects of digestibility measurements of feedstuffs for equines were studied in four Welsh pony geldings consuming four grass-hay diets in a 4 × 4 Latin square design. Diets contained either a low (L), medium (M), high (H), or very high (VH) ADF content (264, 314, 375, or 396 g·kg DM, respectively). Diets were supplemented with minerals, vitamins, and TiO (3.9 g Ti·d). Daily feces excreted were collected quantitatively over 10 consecutive days and analyzed for moisture, ash, ADL, AIA, and titanium (Ti). Minimum duration of total fecal collection (TFC) required for an accurate estimation of apparent organic matter digestibility (OMD) of grass hay was assessed. Based on literature and the calculated cumulative OMD assessed over 10 consecutive days of TFC, a minimum duration of at least 5 consecutive days of fecal collection is recommended for accurate estimation of dry matter digestibility (DMD) and OMD in ponies. The 5-d collection should be preceded by a 14-d adaptation period to allow the animals to adapt to the diets and become accustomed to the collection procedures. Mean fecal recovery over 10 d across diets for ADL, AIA, and Ti was 93.1% (SE 1.9), 98.9% (SE 5.5), and 97.1% (SE 1.8), respectively. Evaluation of CV of mean fecal recoveries obtained by ADL, AIA, and Ti showed that variation in fecal Ti (6.8) and ADL excretion (7.0) was relatively low compared to AIA (12.3). In conclusion, the use of internal ADL and externally supplemented Ti are preferred as markers to be used in digestibility trials in equine fed grass-hay diets.

In- and outdoor reproduction of first generation common sole Solea solea under a natural photothermal regime: Temporal progression of sexual maturation assessed by monitoring plasma steroids and gonadotropin mRNA expression.

Reproduction of many temperate fishes is seasonal and maturation and spawning of gametes are under photothermal control. Reproductive success of first generation (G1) common sole Solea solea in captivity has been low. In this study, the sexual maturation status has been assessed during the prespawning months in G1 sole that were housed (a) outdoor under the natural photoperiod and temperature, or (b) indoor under artificial photothermal induction. Maturation was assessed in male and female G1 broodstock in November as controls, after which the remaining population was divided over two outdoor flow-through tanks placed in a pond and two indoor recirculating aquaculture system (RAS) tanks. Subsequently, maturation status (gonadosomatic index GSI and plasma levels of testosterone T and 17ß-estradiol E2) was assessed in one tank for each condition in January, February and during spawning in early April, while fish in the other tank were not disturbed in achieving reproductive success. Quantitative real-time PCR was performed to determine species-specific gonadotropin mRNA expression in females. Successful G1 spawning and egg fertilisation occurred in all experimental tanks. Gonadal development was similar under both conditions. Higher E2 and T levels were found in indoor housed females. Gonadotropin expression revealed similar profiles between outdoor and indoor housed females. G1 sole could be reproduced in the outdoor tanks under the natural photoperiod and in the indoor tanks under artificial simulation of this regime that includes a potentially crucial chilling period of 2-3 months at 5-7 °C.

The energy and protein value of wheat, maize and blend DDGS for cattle and evaluation of prediction methods.

The chemical composition inclusive amino acids (AAs) and the energy and protein value of three wheat, three maize and seven blend (mainly wheat) dried distillers grains and solubles (DDGS) were determined. The net energy for lactation (NEL) was derived from digestion coefficients obtained with sheep. The digestible protein in the intestines (DVE) and the degraded protein balance (OEB) were determined by nylon bag incubations in the rumen and the intestines of cannulated cows. Additional chemical parameters like acid-detergent insoluble CP (ADICP), protein solubility in water, in borate-phosphate buffer and in pepsin-HCl, in vitro digestibility (cellulase, protease, rumen fluid) and colour scores (L*, a*, b*) were evaluated as potential predictors of the energy and protein value. Compared to wheat DDGS (WDDGS), maize DDGS (MDDGS) had a higher NEL-value (8.49 v. 7.38 MJ/kg DM), a higher DVE-content (216 v. 198 g/kg DM) and a lower OEB-value (14 v. 66 g/kg DM). The higher energy value of MDDGS was mainly due to the higher crude fat (CFA) content (145 v. 76 g/kg DM) and also to better digestible cell-walls, whereas the higher protein value was mainly due to the higher percentage of rumen bypass protein (RBP: 69.8 v. 55.6%). The NEL-value of blend DDGS (BDDGS) was in between that of the pure DDGS-types, whereas its DVE-value was similar to MDDGS. Although lower in CP and total AAs, MDDGS provided a similar amount of essential AAs as the other DDGS-types. Lysine content was most reduced in the production of WDDGS and cysteine in MDDGS. Fat content explained 68.6% of the variation in NEL, with hemicellulose and crude ash as extra explaining variables. The best predictor for RBP as well as for OEB was the protein solubility in pepsin-HCl (R 2=77.3% and 83.5%). Intestinal digestibility of RBP could best be predicted by ADF (R 3=73.6%) and the combination of CFA and NDF could explain 60.2% of the variation in the content of absorbable microbial protein. The availability of AAs could accurately be predicted from the rumen bypass and intestinal digestibility of CP.

Relationship between chemical composition and in situ rumen degradation characteristics of maize silages in dairy cows.

Several in situ studies have been conducted on maize silages to determine the effect of individual factors such as maturity stage, chop length and ensiling of maize crop on the rumen degradation but the information on the relationship between chemical composition and in situ rumen degradation characteristics remains scarce. The objectives of this study were to determine and describe relationships between the chemical composition and the rumen degradation characteristics of dry matter (DM), organic matter (OM), CP, starch and aNDFom (NDF assayed with a heat stable amylase and expressed exclusive of residual ash) of maize silages. In all, 75 maize silage samples were selected, with a broad range in chemical composition and quality parameters. The samples were incubated in the rumen for 2, 4, 8, 16, 32, 72 and 336 h, using the nylon bag technique. Large range was found in the rumen degradable fractions of DM, OM, CP, starch and aNDFom because of the broad range in chemical composition and quality parameters. The new database with in situ rumen degradation characteristics of DM, OM, CP, starch and aNDFom of the maize silages was obtained under uniform experimental conditions; same cows, same incubation protocol and same chemical analysis procedures. Regression equations were developed with significant predictors (P<0.05) describing moderate and weak relationships between the chemical composition and the washout fraction, rumen undegradable fraction, potentially rumen degradable fraction, fractional degradation rate and effective rumen degradable fraction of DM, OM, CP, starch and aNDFom.

Association of bacteriorhodopsin with lipid-impregnated filters. Evidence for fusion of bacteriorhodopsin-containing vesicles with the lipid phase of the filter.

Bacteriorhodopsin vesicles were associated with cellulose-nitrate filters impregnated with a solution of phospholipids in hexadecane. The generation of (photo)potentials upon illumination of the filter was studied in the absence and presence of ionophores, phospholipase A2, EDTA or polyene antibiotics. From these experiments the following conclusions are drawn. 1. Upon illumination of the filter, bacteriorhodopsin pumps protons into aqueous compartments located in the filter. 2. These aqueous compartments possibly do not originate from the compartments enclosed by the bacteriorhodopsin vesicles. Evidence is obtained that aqueous compartments are present in the surface layers of the lipid-impregnated filters. 3. The results are explained most easily by a mechanism, whereby fusion occurs between the vesicles and the lipids of the filter.

Light-induced pH changes inside bacteriorhodopsin vesicles as measured by 31 P NMR.

31P NMR has been used to measure light-induced pH changes inside bacteriorhodopsin vesicles containing entrapped sodium glucose-6-phosphate. Reversible light-induced pH changes were observed at various pH values. The results indicate that our vesicle preparations were not homogeneous with respect to the generation of pH gradients.

A molecular basis for an irreversible thermodynamic description on non-electrolyte permeation through lipid bilayers.

The non-electrolyte permeability of liposomal membranes has been investigated according to the concepts of irreversible thermodynamics. A strong interaction between the permeation of solute and water was observed. This solute-solvent interaction can be fully described by assuming that a number of water molecules will copermeate with each molecule of solute. This number of copermeating water molecules is independent of the nature of the permeant and of temperature, but depends on the osmotic concentration of impermeants inside the liposomes.

Non-electrolyte permeability as a tool for studying membrane fluidity.

1. The reflection coefficient for the permeation of thiourea through bilayers of phosphatidylcholine is a function of the fatty-acid composition of the lipid molecules. By means of these reflection coefficients an index for membrane fluidity has been given to each of those lipids, relative to that of egg phosphatidylcholine. 2. The maximum number of water molecules that can copermeate with each molecule of solute by means of solute-solvent interaction is a function of the packing of the lipid molecules in the bilayer. This parameter has been used in this paper for characterizing the fluidity of cholesterol-containing membranes and for membranes with their lipids in the gel state.

The effect of cholesterol incorporation on the temperature dependence of water permeation through liposomal membranes prepared from phosphatidylcholines.

The permeation of water through liposomal membranes composed of phosphatidylcholine plus varying amounts of cholesterol was studied as a function of temperature. 1. Increasing amounts of cholesterol caused a gradual disappearance of the abrupt change in the rate of water permeation near the gel to liquid-crystalline phase transition temperature of dipalmitoylphosphatidylcholine and dimyristoylphosphatidylcholine liposomes. At cholesterol concentrations above about 30 mol % there was no longer a discontinuity in the rate of water permeation. 2. The incorporation of cholesterol produces a steep change in the activation energy of the water permeation above the transition temperature of the saturated lecithin occurring at about 15 mol % of cholesterol. Below the transition temperature there was a gradual decrease in the activation energy of the water permeation in the region of 0 to 33 mol % of cholesterol. 3. In systems containing unsaturated phosphatidylcholines cholesterol also enhanced the activation energy of the water permeation although to a lesser extent. The results indicate that the position of the cis-double bond in the fatty acid chain is very important in this respect. 4. In systems in which cholesterol increased the temperature dependence of the water permeation there is also an enhancement of the temperature dependence of the isotonic glycerol and erythritol swelling by the same number of kcal/mol.

Effect of the gel to liquid crystalline phase transition on the osmotic behaviour of phosphatidylcholine liposomes.

Aspects of osmotic properties of liposomes, prepared from synthetic lecithin, above, at and below the gel to liquid crystalline phase transition temperature are described. The experiments show that liposomal membranes with their lipids in the gel state are still permeable to water. The rate of water permeation changes drastically on passing the transition temperature. The water permeation has activation energies of 9.5 +/- 1.28 and 26.4 +/- 0.9 kcal/mol above and below the transition temperature, respectively, indicating that the diffusion processes take place by different mechanisms. With respect to the barrier properties of the liposomes in the vicinity of the transition temperature, the following conclusions can be made. (1) Studying the osmotic shrinkage of liposomes at a fixed temperature near the transition point, the experiments indicate that dimyristoyl phosphatidylcholine membranes are highly permeable to glucose under these conditions, where liquid and solid domains co-exist. Under the same conditions the osmotic experiments did not indicate a strong increase in glucose permeability of dipalmitoyl phosphatidylcholine membranes as compared to the situation above and below the transition temperature. (2) On the other hand, perturbations of the phase equilibrium by temperature varations resulted in a marked increase of the glucose permeation through dipalmitoyl phosphatidylcholine bilayers. Once a new phase equilibrium of liquid and solid regions is established the permeation rate of glucose is much less.