The interaction between protein and phytate was investigated in vitro using proteins extracted from five common feedstuffs and from casein. The appearance of naturally present soluble protein-phytate complexes in the feedstuffs, the formation of complexes at different pHs, and the degradation of these complexes by pepsin and/or phytase were studied. Complexes of soluble proteins and phytate in the extracts appeared in small amounts only, with the possible exception of rice pollards. Most proteins dissolved almost completely at pH 2, but not after addition of phytate. Phytase prevented precipitation of protein with phytate. Pepsin could release protein from a precipitate, but the rate of release was increased by phytase. Protein was released faster from a protein-phytate complex when phytase was added, but phytase did not hydrolyze protein. Protein was released from the complex and degraded when both pepsin and phytase were added. It appears that protein-phytate complexes are mainly formed at low pH, as occurs in the stomach of animals. Phytase prevented the formation of the complexes and aided in dissolving them at a faster rate. This might positively affect protein digestibility in animals.

A (31)P NMR method for quantitative determination of inositol phosphates in simple incubation samples of sodium phytate and Aspergillus niger phytase and in different types of complex samples, such as diets, digesta, and feces, is described. The inositol phosphates in complex samples were extracted with HCl, concentrated, and purified using freeze-drying and filtration and subsequently determined at pH 12.6 in aqueous solution using a (31)P NMR method. The (31)P NMR technique has as its main advantages over the HPLC techniques that it does not necessitate standards that may cause background matrix effects and that the spectra of inositol phosphates and orthophosphate appear in the same run without further sampling errors. The results of inositol hexaphosphate analysis with HPLC can be confirmed by this (31)P NMR method. Contents of inositol tetra-, tri-, di-, and monophosphate in the biological samples appear to be quantitatively not important. The (31)P NMR method can be applied for use in animal nutrition in general and studies of using phytase in diets for farm animals in particular, by measuring the content of inositol phosphates in feed ingredients, complete feeds, ileal contents, and feces of pigs and poultry.

The objective of this study was to investigate the effects of increasing maturity of whole-plant corn at harvest on CH4 emissions by dairy cows consuming corn silage (CS) based diets. Whole-plant corn was harvested at a very early [25% dry matter (DM); CS25], early (28% DM; CS28), medium (32% DM; CS32), and late (40% DM; CS40) stage of maturity. In a randomized block design, 28 lactating Holstein-Friesian dairy cows, of which 8 were fitted with rumen cannula, received 1 of 4 dietary treatments designated as T25, T28, T32, and T40 to reflect the DM contents at harvest. Treatments consisted of (DM basis) 75% CS, 20% concentrate, and 5% wheat straw. Feed intake, digestibility, milk production and composition, energy and N balance, and CH4 production were measured during a 5-d period in climate respiration chambers after an adaptation to the diet for 12 d. Corn silage starch content varied between 275 (CS25) and 385 (CS40) g/kg of DM. Treatments did not affect DM intake (DMI), milk yield, or milk contents. In situ ruminal fractional degradation rate of starch decreased linearly from 0.098 to 0.059/h as maturity increased from CS25 to CS40. Apparent total-tract digestibility of DM, organic matter, crude protein, neutral detergent fiber, crude fat, starch, and gross energy (GE) decreased linearly with maturity. Treatments did not affect ruminal pH, volatile fatty acids, and ammonia concentrations, and volatile fatty acids molar proportions. The concentration of C18:3n-3 in milk fat decreased linearly, and the concentration of C18:2n-6 and the n-6:n-3 ratio increased linearly with maturity. A quadratic response occurred for the total saturated fatty acid concentration and total monounsaturated fatty acid concentration in milk fat. Methane production relative to DMI (21.7, 23.0, 21.0, and 20.1g/kg) and relative to GE intake (0.063, 0.067, 0.063, and 0.060 MJ/MJ; values for T25, T28, T32, and T40, respectively) decreased linearly with maturity. Also, CH4 emission relative to fat- and protein-corrected milk tended to decrease linearly with maturity (13.0, 13.4, 13.2, and 12.1g/kg of fat- and protein-corrected milk, for T25, T28, T32, and T40, respectively). Intake of GE and metabolizable energy, and energy retained, all expressed per unit of metabolic body weight, did not differ among treatments. Nitrogen intake, N use efficiency (milk N/N intake), and N balance were not influenced by treatments. Increasing maturity of whole-plant corn at harvest may offer an effective strategy to decrease CH4 losses with feeding CS without negatively affecting cow performance.

A modified rinsing method for the in situ technique was developed to separate, isolate and characterise the soluble (S), the insoluble washout (W-S) and the non-washout fractions (D 1 U) within one procedure. For non-incubated bags (t 5 0 h), this method was compared with the conventional, Combined Fractionation (CF) method that measures the D 1 U and S fractions in separate steps and subsequently calculates the W-S fraction. The modified method was based on rinsing of nylon bags in a closed vessel containing a buffer solution (pH 6.2) during 1 h, where shaking speeds of 40, 100, and 160 strokes per minutes (spm) were evaluated, and tested for six feed ingredients (faba beans, maize, oats, peas, soya beans and wheat) and four forages (two ryegrass silages and two maize silages). The average recoveries as the sum of all fractions were 0.972 6 0.041 for N and 0.990 6 0.050 for starch (mean 6 s.d.). The mean W-S fraction increased with increasing shaking speed and varied between 0.017 (N) and 0.083 (starch) at 40 spm and 0.078 (N) and 0.303 (starch) at 160 spm, respectively. For ryegrass silages, the W-S fraction was absent at all shaking speeds, but was present in the CF method. The modified method, in particular at 40 and 100 spm, reduced the loss of small particles during rinsing, resulting in lower W-S and higher D 1 U fractions for N and starch compared with the CF method. For soya beans and ryegrass silage, the modified method reduced the S fraction of N compared with the CF method. The results obtained at 160 spm showed the best comparison with those from the CF method. The W-S fraction of the feedstuff obtained at 160 spm contained mainly particles smaller than 40 mm (0.908 6 0.086). In most feedstuff, starch was the most abundant chemical component in the W-S fraction and its content (726 6 75 g/kg DM) was higher than in the D 1 U fraction (405 6 177 g/kg DM). Alkaline-soluble proteins were the dominant N-containing components in the W-S fraction of dry feed ingredients and its relative content (0.79 6 0.18 of total N in W-S) was higher than in the D 1 U fraction (0.59 6 0.07 of total N in D 1 U) for all feedstuff except maize. The molecular weight distribution of the alkaline-soluble proteins differed between the W-S and the D 1 U fractions of all dry feed ingredients, except soya beans and wheat.

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The objective of this experiment was to evaluate interaction effects between stage of maturity and nitrogen (N) fertilisation rate on rumen degradation characteristics determined with nylon bag incubations of ryegrass herbages and ryegrass silage. Grass herbage (n = 4) was cut after 3 or 5 weeks of regrowth and received a low (20 kg N/ha) or a high (90 kg N/ha) fertilisation rate. Grass silage (n = 6) received a low (65 kg N/ha) or high (150 kg N/ha) fertilisation rate and was harvested at early (~2000 kg DM/ha), mid (harvested 13 days later), or late (harvested 34 days later) maturity stage and ensiled in big bales. All grasses were incubated in the rumen of three lactating rumen-cannulated Holstein Friesian cows. Rumen degradation characteristics of organic matter (OM), N and neutral detergent fibre (NDF) and the extent of effective degradation (ED) were evaluated. In grass herbage, NDF content varied between 390 and 454 g/kg DM and N content between 12.1 and 25.8 g/kg DM. In grass silage, NDF content varied between 438 and 593 g/kg DM and N content between 13.4 and 34.8 g/kg DM. In general, rumen degradation of grass herbage and grass silage decreased with increased maturity, and increased with increased fertilisation rate. Significant interaction between maturity and fertilisation rate was observed for ED of OM, N and NDF, except for ED of N in grass herbage. These results indicate that the effect of the rate of N fertilisation on degradation of nutrients in the rumen of dairy cattle and on nutritional value depends on the grass maturity stage.