Dietary fiber components: relationship to the rate and extent of ruminal digestion

A mathematical model can serve as a useful reference for describing the mechanisms involved in digestion and for discussing the factors that influence the rate and extent of ruminal digestion. Ruminal digestion can be divided into four components: digestion rate, digestion lag, potential extent of digestion, and passage rate. Each component affects the apparent extent of digestion in a distinct manner and is influenced by separate factors. Digestion rate is directly related to apparent extent of digestion. It is not influenced by chemical entities presently being measured, but may be related to the morphological, crystalline, or physical nature of fiber. It may also be influenced by factors that inhibit or stimulate ruman microbial growth and their fiber-degrading enzymes. Digestion lag is inversely related to apparent extent of digestion; however, factors influencing it are poorly defined. The may include factors affecting microbial populations and their attachment to fiber prior to digestion; or the digestion lag may be related to the chemical or physical alteration of fiber that must occur before digestion can begin. The potential extent of digestion is directly related to apparent extent of digestion and is influenced by plant fiber composition, primarily. Lignin, and possibly silica, functions to limit the potential extent of digestion. Rate of passage essentially competes with rate of digestion for fiber particles as they pass through the rumen; therefore it is inversely related to the apparent extent of digestion. Passage rate is associated with feed intake level and particle size, although other factors such as type of diet and animal physiology may be important.     

In vitro and in vivo phosphorylation of polypeptides in plasma membrane and tonoplast-enriched fractions from barley roots

Phosphorylation of polypeptides in membrane fractions from barley (Hordeum vulgare L. cv CM 72) roots was compared in in vitro and in vivo assays to assess the potential role of protein kinases in modification of membrane transport. Membrane fractions enriched in endoplasmic reticulum, tonoplast, and plasma membrane were isolated using sucrose gradients and the membrane polypeptides separated using sodium dodecyl sulfate polyacrylamide gel electrophoresis. When the membrane fractions were incubated with γ-[³²P]ATP, phosphorylation occurred almost exclusively in the plasma membrane fraction. Phosphorylation of a band at 38 kilodaltons increased as the concentration of Mg²⁺ was decreased from millimolar to micromolar levels. Phosphorylation of bands at 125, 86, 58, 46, and 28 kilodaltons required millimolar Mg²⁺ concentrations and was greatly enhanced by Ca²⁺. When roots of intact plants were labeled with [³²P]orthophosphate, polypeptides at approximately 135, 116, 90, 46 to 53, 32, 28, and 19 kilodaltons were labeled in the plasma membrane fraction and polypeptides at approximately 73, 66, and 48 kilodaltons were labeled in the tonoplast fraction. Treatment of the roots of intact plants with 150 millimolar NaCl resulted in increased phosphorylation of some polypeptides while treatment with 100 mm NaCl had no effect.     

On the two-compartment model for estimating the rate and extent of feed degradation in the rumen

An analysis of the compartmental scheme used to determine the rate and extent of ruminal degradation of feeds is presented. Attention is given to the kinetic representation of the degradation of the potentially degradable fraction. Changing the kinetic order of the rate, and introducing indigestible substrate inhibition and microbial activity into its representation, are investigated. This leads to response functions such as the Gompertz and logistic for describing the cumulative disappearance of potentially degradable substrate during in-sacco and in-vitro incubation.     

Effect of maize silage to grass silage ratio and feed particle size on ruminal fermentation in vitro

The effect of the forage source on ruminal fermentation in vitro was investigated for fine (F) and coarse (C) milled diets, using a modified Hohenheim gas production test and a semi-continuous rumen simulation technique (Rusitec). It was hypothesised that the replacement of maize silage by grass silage might lead to associative effects and that interactions related to particle size variation could occur. Five diets with a maize silage to grass silage ratio of 100 : 0, 79 : 21, 52 : 48, 24 : 76 and 0 : 100 differed in their content of CP and carbohydrate fractions, as well as digestible crude nutrients, derived from a digestibility trial with wether sheep. For in vitro investigations, the diets were ground to pass a sieve of either 1 mm (F) or 4 mm (C) perforation. Cumulative gas production was recorded during 93 h of incubation and its capacity decreased with increasing proportion of grass silage in the diet. Across all diets, gas production was delayed in C treatments compared with F treatments. Degradation of crude nutrients and detergent fibre fractions was determined in a Rusitec system. Daily amounts of NH3-N and short-chain fatty acids (SCFA) were measured in the effluent. Degradation of organic matter (OM) and fibre fractions, as well as amounts of NH3-N, increased with stepwise replacement of maize silage by grass silage. Degradability of CP was unaffected by diet composition, as well as total SCFA production. In contrast to the results of the gas production test, degradation of OM and CP was higher in C than in F treatments, accompanied by higher amounts of NH3-N and SCFA. Interactions of silage ratio and particle size were rare. It was concluded that the stepwise replacement of maize silage by grass silage might lead to a linear response of most fermentation characteristics in vitro. This linear effect was also supported by total tract digestibility data. However, further investigations with silages of variable quality seem to be necessary.     

In vitro genotoxicity data of nanomaterials compared to carcinogenic potency of inorganic substances after inhalational exposure

Literature data of epidemiological studies, carcinogenicity studies and in vitro studies on inorganic substances were surveyed with the aim to determine sensitivity and specificity of in vitro tests of nanomaterials. Asbestos, quartz and chromium and cadmium compounds were assigned to classes of highest carcinogenic potency. After 20 years of occupational exposure to long-term average concentrations of 0.5mg/m(3) of these dusts - or to even lower concentrations - an epidemiologically detectable increased lung cancer risk has to be expected. In contrast, diesel engine emissions, some nickel species and "ultrafine" versions (nanomaterials) of titanium dioxide and carbon black were also carcinogenic in inhalation studies, but show varied epidemiological results. The high frequency of lung cancer in the male general population due to cigarette smoking hampers unequivocal detection of occupationally caused lung cancer risks. Based on the experience from the inhalation studies, workers had to be exposed to long-term concentrations of 1mg/m(3) or more to identify epidemiologically a clear cause-and-effect relationship for a specific substance of intermediate potency. Respirable granular biodurable particles without known significant specific toxicity with primary particle sizes of more than 1μm have also shown carcinogenicity in rats. Their potency was even lower; and partially results after instillation rather than inhalation are available. Nearly all types of nanomaterials and control dusts used in the in vitro assays showed genotoxic effects in cell cultures (e.g., CoCr particles, diesel soot, SiO(2) crystalline and amorphous, TiO(2), carbon black), but not consistently in all studies; overall, the proportion of positive results was about 50%. No clear correlation of the probability of a positive in vitro test with particle properties was seen. I recommend trying and calibrating a sensitive in vitro model (e.g., micronucleus assay) against the described rank order of carcinogenic potency by testing a series of inorganic substances.     

In vitro digestion and fermentation characteristics of canola co-products simulate their digestion in the pig intestine

Canola co-products are sources of amino acid and energy in pig feeds, but their fermentation characteristics in the pig intestine are unknown. Thus, we determined the in vitro fermentation characteristics of the canola co-products Brassica juncea solvent-extracted canola meal (JSECM), Brassica napus solvent-extracted canola meal (NSECM), B. napus expeller-pressed canola meal (NEPCM) and B. napus cold-pressed canola cake (NCPCC) in comparison with soybean meal (SBM). Samples were hydrolysed in two steps using pepsin and pancreatin. Subsequently, residues were incubated in a buffer solution with fresh pig faeces as inocula for 72 h to measure gas production. Concentration of volatile fatty acids (VFA) per gram of dry matter (DM) of feedstuff was measured in fermented solutions. Apparent ileal digestibility (AID) and apparent hindgut fermentation (AHF) of gross energy (GE) for feedstuffs were obtained from pigs fed the same feedstuffs. On DM basis, SBM, JSECM, NSECM, NEPCM and NCPCC contained 15, 19, 22, 117 and 231 g/kg ether extract; and 85, 223, 306, 208 and 176 g/kg NDF, respectively. In vitro digestibility of DM (IVDDM) of SBM (82.3%) was greater (P<0.05) than that of JSECM (68.5%), NSECM (63.4%), NEPCM (67.5%) or NCPCC (69.8%). The JSECM had greater (P<0.05) IVDDM than NSECM. The IVDDM for NSECM was lower (P<0.05) than that for NEPCM, which was lower (P<0.05) than that for NCPCC. Similarly, AID of GE was greatest for SBM followed by NCPCC, JSECM, NEPCM and then NSECM. Total VFA production for SBM (0.73 mmol/g) was lower (P<0.05) than that of JSECM (1.38 mmol/g) or NSECM (1.05 mmol/g), but not different from that of NEPCM (0.80 mmol/g) and NCPCC (0.62 mmol/g). Total VFA production of JSECM was greater (P<0.05) than that of NSECM. Total VFA production of NSECM was greater (P<0.05) than that of NEPCM or NCPCC, which differed (P<0.05). The ranking of feedstuffs for total VFA production was similar to AHF of GE. In conclusion, in vitro fermentation characteristics of canola co-products and SBM simulated their fermentation in the small and large intestine of pigs, respectively. The 30% greater VFA production for JSECM than NSECM due to lower lignified fibre of JSECM indicates that fermentation characteristics differ between canola species. The NSECM had the highest fermentability followed by NEPCM and then NCPCC, indicating that fat in canola co-products can limit their fermentability in the hindgut.     

In vitro rumen fermentation of soluble and non-soluble polymeric carbohydrates in relation to ruminal acidosis

The end-products of dietary carbohydrate fermentation catalysed by rumen microflora can serve as the primary source of energy for ruminants. However, ruminants provided with continuous carbohydrate-containing feed can develop a metabolic disorder called “acidosis”. We have evaluated the fermentation pattern of both soluble monomeric and non-soluble polymeric carbohydrates in the rumen in in vitro fermentation trials. We found that acidosis could occur within 6 h of incubation in the rumen culture fermenting sugars and starch. The formation of lactic acid and acetic acid, either alone or in mixture with ethanol, accounted for high build-up of acid in the rumen. Acidosis resulted even when only 20% of a normal daily feed load for all soluble and non-soluble carbohydrates was provided. DNA-based microbial analysis revealed that Prevotella was the dominant microbial species present in the rumen fluid.     

Susceptibility and resistance of ruminal bacteria to antimicrobial feed additives

Susceptibility and resistance of ruminal bacterial species to avoparcin, narasin, salinomycin, thiopeptin, tylosin, virginiamycin, and two new ionophore antibiotics, RO22-6924/004 and RO21-6447/009, were determined. Generally, antimicrobial compounds were inhibitory to gram-positive bacteria and those bacteria that have gram-positive-like cell wall structure. MICs ranged from 0.09 to 24.0 micrograms/ml. Gram-negative bacteria were resistant at the highest concentration tested (48.0 micrograms/ml). On the basis of their fermentation products, ruminal bacteria that produce lactic acid, butyric acid, formic acid, or hydrogen were susceptible and bacteria that produce succinic acid or ferment lactic acid were resistant to the antimicrobial compounds. Selenomonas ruminantium was the only major lactic acid-producing bacteria resistant to all the antimicrobial compounds tested. Avoparcin and tylosin appeared to be less inhibitory (MIC greater than 6.0 micrograms/ml) than the other compounds to the two major lactic acid-producing bacteria, Streptococcus bovis and Lactobacillus sp. Ionophore compounds seemed to be more inhibitory (MIC, 0.09 to 1.50 micrograms/ml) than nonionophore compounds (MIC, 0.75 to 12.0 micrograms/ml) to the major butyric acid-producing bacteria. Treponema bryantii, an anaerobic rumen spirochete, was less sensitive to virginiamycin than to the other antimicrobial compounds. Ionophore compounds were generally bacteriostatic, and nonionophore compounds were bactericidal. The specific growth rate of Bacteroides ruminicola was reduced by all the antimicrobial compounds except avoparcin. The antibacterial spectra of the feed additives were remarkably similar, and it appears that MICs may not be good indicators of the potency of the compounds in altering ruminal fermentation characteristics.     

Acclimation of Rubisco specificity factor to drought in tobacco: discrepancies between in vitro and in vivo estimations

In studies about the photosynthesis response to environmental stresses, such as drought, the Rubisco specificity factor (tau) is assumed to be constant or derived indirectly from gas exchange measurements. However, an analysis of the acclimation of tau to drought using in vitro determinations is lacking. The aim of the present work was to analyse the acclimation of tau to different drought intensities in tobacco (Nicotiana tabacum L.). Potted tobacco plants were subjected to three different water regimes (100%, 40%, and 15% of field capacity) and new leaves were allowed to develop. When acclimated leaves were fully developed, they were sampled for gas exchange and chlorophyll fluorescence measurements, as well as for the in vitro analysis of Rubisco kinetic properties. Relative water content and gas exchange decreased with increasing water shortage. The apparent Rubisco specificity factor as estimated in vivo by gas exchange decreased with water stress. However, in vitro estimates of tau were identical among treatments, as were Rubisco specific initial activity and activation state. The reasons for the observed discrepancy between in vitro and in vivo estimates are profusely discussed. It is suggested that the Rubisco specificity factor does not acclimate to water stress in the short term (weeks or months) in tobacco, and the validity of the so-called Laisk gas exchange method to estimate tau under drought is questioned.     

In vivo and in vitro estimations of the direct effect of estrogen on rat hepatocytes tested by the changes in the unusual estrogen-binding protein content

The direct effect of estradiol (E2) on the hepatocytes of mature male rats has been examined by measuring the changes in the unusual estrogen-binding protein (UEBP) content and parallel measuring the level of liver estrogen receptors (ER). The content of UEBP (NUEBP) and ER (NER) in the liver were determined using the quantitative methods for differential specific determination of the E2-binding sites of these proteins. It has been shown that the administration of E2 in vivo induced a considerable decrease in hepatic NUEBP not only in intact males, but also in hypophysectomized males during the initial period after the operation (when the content of hepatic ER was still high) and produced no effect in hypophysectomized males during the later period (when liver ER were depleted). Repeated administration of human growth hormone (hGH) (twice a day) resulted in a considerable increase in NER in hypophysectomized males and restored the sensitivity to the subsequent inhibitory effect of E2 on UEBP. We also used rat hepatocytes after a 4-day primary culturing. These cells had a stable morpho-functional status, high ER level, and sex-differentiated UEBP content. Culturing of mature male rat hepatocytes in the medium containing E2 at concentrations close to physiological levels (10(-10)-10(-7) M) decreased NUEBP in a dose-dependent manner. Hexestrol (10(-7) M) but not cholesterol (10(-5) M) also exhibited a direct effect on NUEBP in cultured rat hepatocytes. The effect of E2 was reversible: statistically significant increase in NUEBP was observed 3 days after 10(-9) M E2 had been removed from the culturing medium. It was concluded that hepatocytes may be a primary target for E2 under physiological conditions and that GH may modulate the direct effect of E2 at the hepatic level by modifying the content of liver ER.     

In vivo measurement-based estimations of the moment arm in the human tibialis anterior muscle-tendon unit

The aim of this study was to estimate the moment arm of human tibialis anterior (TA) muscle-tendon unit at rest and during isometric dorsiflexion maximum voluntary contraction (MVC) from in vivo sagittal-plane magnetic resonance (MR) and ultrasound scans. Two methods were employed, both of them based on the assumption that the ankle joint complex and TA muscle-tendon unit operate in the sagittal plane. Using method A, moment arms were obtained from MR scans of the foot by measuring the perpendicular distance between a moving centre of rotation in the talo-crural joint and the TA tendon action line. Using method B, moment arms were calculated from the ratio of TA tendon displacement, which was estimated from a planimetric muscle model using pennation angles and muscle thickness measured by ultrasonography, to the tibial rotation around the talus, which was measured from the foot MR scans. Using either of the two methods at rest, the estimated TA moment arm decreased from approximately 4.5 to approximately 2.9 cm in the transition from dorsiflexion to plantarflexion. Using method A, moment arms during MVC were larger by 0.9-1.5 cm (33-44%, P < 0.01) than the respective resting estimations. In contrast, no difference (P > 0.05) was found between the resting and MVC moment arm estimations of method B. Limitations in the oversimplified musculoskeletal model used raise questions for the validity of both method estimations.     

In vitro ruminal fermentation of organic acids common in forage.

Mixed rumen bacteria from cows fed either timothy hay or a 60% concentrate were incubated with 7.5 mM citrate, trans-aconitate, malate, malonate, quinate, and shikimate. Citrate, trans-aconitate, and malate were fermented at faster rates than malonate, quinate, and shikimate. Acetate was the primary fermentation product for all six acids. Quinate and shikimate fermentations gave rist to butyrate, whereas malate and malonate produced significant amounts of propionic acid. High-pressure liquid chromatography of fermentation products from trans-aconitate incubations revealed a compound that was subsequently identified as tricarballylate. As much as 40% of the trans-aconitate acid was converted to tricarballylate, and tricarballylate was fermented slowly. The slow rate of tricarballylate metabolism by mixed rumen bacteria and its potential as a magnesium chelator suggest that tricarballylate formation could be an important factor in the hypomagnesemia that leads to grass tetany.     

Ability of six different lipoprotein fractions to regulate the rate of hepatic cholesterogenesis in vivo.

Two in vivo assay procedures were used to study the inhibitory activity of cholesterol carried in three intestinal lymph and three serum lipoprotein fractions on the rate of cholesterol synthesis in the liver. In the first preparation, different lipoproteins were injected intravenously as a bolus into rats at the mid-light phase of the diurnal light cycle, following which they were killed 12 hours later in the mid-dark phase of the cycle. Using this assay, three intestinal lymph lipoprotein fractions of varying Sf values all produced a similar degree of inhibition which averaged approximately 11%/mg of cholesterol injected. The serum lipoprotein fractions caused only about one-third this amount of inhibition. Detailed analysis of events occurring within the liver during this 12-hour assay period revealed that there were marked differences in the rate of net cholesterol uptake into the liver and in the rate of new removal of cholesterol esters from the liver following injection of each of these different lipoprotein fractions. The amount of inhibition of sterol synthesis produced by any fraction was proportional to the product of the incremental increase in hepatic cholesterol ester content and the time over which this increase in esters occurred. In the second type of assay where the lipoprotein fractions were administered to the animals as a continuous intravenous infusion over 24 hours the largest increase in hepatic cholesterol ester content and the greatest inhibition of cholesterol synthesis was found with intestinal lipoproteins having Sf values larger than 8000. Intestinal lipoprotein fractions with lower Sf values and all serum lipoprotein fractions were significantly less effective in bringing about an increase in hepatic cholesterol ester content and in producing inhibition of cholesterol synthesis by the liver. These studies emphasize the primary role of cholesterol carried in lipoproteins of intestinal origin in regulating hepatic sterol synthesis. The inhibitory activity of these fractions appears to correlate with the ability of these lipoproteins to bring about a maximal increase in hepatic cholesterol ester content which, in turn, appears to relate to the capacity of these fractions to transfer cholesterol rapidly into the hepatocyte while, at the same time, slowing the rate of cholesterol mobilization from the liver.     

Effect of rare earth elements on in vitro rumen microbial fermentation and feed digestion

The objectives of this study were to investigate the effects of rare earth elements (REEs) on in vitro rumen fermentation, gas production, microbial protein synthesis and nutrient digestion using in vitro batch culture and continuous culture technique. A mixture of REE containing (g/kg) 380 g of LaCI₃·6H₂O, 521 g of CeCI₃·6H₂O, 30 g of PrCI₃·6H₂O and 69 g chlorides of other light REEs. The experimental diet consisted of 885 g/kg barley grain, 84 g/kg barley silage and 31 g/kg supplement (dry matter (DM) basis). Diet supplemented with different dosages of REE (control, no additional REE; low, 400 mg/kg REE; and high, 800 mg/kg REE, DM basis) were incubated for 4, 8, 14 and 24 h in diluted rumen fluid. At the end of 24 h of incubation, gas production and concentration of volatile fatty acid (VFA) linearly increased with increasing REE supplementation; whereas, influence of REE supplementation on VFA profile was marginal. Dry matter disappearance was not affected (P>0.10). Six dual-flow continuous culture fermenters were used in a replicated 3 × 3 Latin square with same treatments and same diet used in the batch culture. Mean ruminal pH (5.71) and total VFA (93.6 mM) concentration were not affected by supplementation of REE. The molar proportion (mol/100 mol) of acetate (39.1) and propionate (50.5) was similar among the treatments. However, the proportion (mol/100 mol) of butyrate was higher with the high REE (6.6) than with low REE (5.3) or the control (5.8). Ruminal true digestibilities of organic matter (OM) (0.785, 0.811 and 0.828), acid detergent fibre (0.360, 0.431 and 0.432) and crude protein (0.496, 0.590 and 0.589) for control, low and high REE, respectively, linearly increased with increasing REE supplementation, whereas, the increase in ruminal digestibility from low to high dosage of REE was minimal. Microbial nitrogen (N) production (g/day) and microbial efficiency (g N/kg of truly fermented OM) were not affected by treatments. Improvement of ruminal digestibility of OM due to REE supplementation was attributed to the increase in digestibility of fibre and degradability of protein. The results suggest that REE supplementation improved ruminal fibrolytic and proteolytic activities.