Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “DIGESTION”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 523 records · Page 29Linked to original sources
PubMed2000

Quantifying the digestibility of dietary protein.

The current recommendation, when calculating a protein digestibility-corrected amino acid score, is to determine the digestibility of a dietary protein across the entire digestive tract, using the rat as a model animal for humans. This fecal digestibility value is subsequently corrected for endogenous contributions of protein using a metabolic nitrogen value determined by feeding rats a protein-free diet. The limitations inherent with this method are well recognized, however, and determining the digestibility of a dietary protein to the end of the small intestine is the preferred alternative. Unlike the fecal digestibility assay, which has only one basic methodology, ileal digestibility values can be determined in a number of ways. We discuss the various methods available for determining ileal digestibility values and compare results obtained for dietary proteins using both fecal and ileal digestibility assays. The relative value of using individual amino acid digestibility values as opposed to nitrogen digestibility values is reviewed. In addition, we address issues surrounding measurement of endogenous nitrogen flows, and in particular, the relative merits of determining "true" versus "real" digestibility values.

Adult↗
PubMed2003

Effect of enzyme addition on the digestibilities of cell wall polysaccharides and oligosaccharides from whole, dehulled, and ethanol-extracted white lupins in chickens.

Three experiments were conducted to examine the effects of a commercial enzyme preparation on chicks performance and digestibilities of nonstarch polysaccharides (NSP), raffinose (R), stachyose (S), and total oligosaccharides (O) in diets containing whole and dehulled lupin and ethanol-extracted dehulled lupin meal. Ethanol extract was also used to produce a rich oligosaccharide fraction. In the first experiment, the dehulling treatment and the addition of enzyme to the diet improved (P < 0.05) weight gain (24 and 15%), feed-to-gain ratio (13 and 9%), dry matter retention (32 and 8%), apparent protein digestibility (6 and 3%) and ileal digestibilities of raffinose (19 and 119%), stachyose (85 and 204%), and total oligosaccharides (68 and 178%), respectively. In addition, enzyme treatment improved (P < 0.05) excreta digestibility values for NSP (from -1.7 to 5.5%), R (from 64.6 to 92.6%), S (from 48.8 to 82.4%), and O (from 55.8 to 83.5%). In the second experiment, extraction of the ethanol soluble components from dehulled lupin decreased (P < 0.0001) weight gain (51%), and increased (P < 0.0001) feed consumption (34%), feed-to-gain ratio (32%), relative gizzard weight (14%), and the relative ceca length (20%). The addition of the isolated fraction of oligosacharides to the corn-extracted lupin diet (7.5 and 15%) yielded performance values similar to those obtained with the unextracted dehulled lupin. The addition of enzymes to the diets significantly improved the weight gain (11%) and feed to gain ratio (6%), and decreased relative gizzard weight (12%) and relative ceca length (7%). Ileal digestibilities of R, S, and O were considerably lower than corresponding excreta digestibilities. The excreta digestibility of NSP and excreta and ileal digestibilities of R, S, and O were lower (P < 0.05) in chicks fed the lupin diet containing the high concentration of extract compared with those fed the same diet containing the lower concentration of extract. Enzymes, when added to the diets, increased (P < 0.05) the digestibilities of R, S, and O with the relative effects being much greater for ileal than excreta samples. In the third experiment, the ethanol extract (7.5 and 15%) added to a corn-soybean diet improved weight gain, feed consumption, and feed-to-gain ratio by 19, 13, and 6%, respectively. Ileal digestibilities of R, S, and O were low (< 45%), especially in chicks fed the diets containing the high amount of added oligosaccharide fraction. There was no effect of enzyme addition on the performance parameters. However, supplemental dietary enzymes improved (P < 0.05) ileal and excreta digestibilities of oligosaccharides from 15.6 and 68.6% to 52.3 and 84.8%, respectively. In summary, dehulling greatly improved the nutritional value of lupin, while removal of the ethanol soluble materials, including raffinose and stachyose considerably decreased its value. The oligosaccharide fraction does not seem to have an antinutritive effect and the addition of enzymes increased the digestibilities of NSP, raffinose, and stachyose but did not significantly improve chick performance.

Animal Feed↗
PubMed2003

Effects of microflora status, dietary bile salts and guar gum on lipid digestibility, intestinal bile salts, and histomorphology in broiler chickens.

The aim of the present study was to reexamine the effects of intestinal viscosity, microflora, and bile salts, and their interactions in order to clarify the mechanisms that explain the effect of intestinal viscosity on lipid digestibility, especially those that could involve microflora. Effects of intestinal viscosity, microflora status, and dietary bile salts on lipid digestibility, intestinal bile salts, and intestinal histomorphology were tested in a 2 x 2 x 2 factorial experiment. The effect of microflora status was examined by comparing conventional chickens to chickens with limited microflora obtained by rearing birds in sterilized conditions. Viscosity and dietary bile salts were tested with guar gum (0 or 0.5% in diets) and sodium taurocholate (0 or 0.3% in diets), respectively. Guar gum was autoclaved and added to the gamma-irradiated diets by mixing inside the sterile isolators. The intestinal concentration of lactic acid and cecal concentration of short-chain fatty acids were both very low in birds with limited microflora compared to conventional birds (P = 0.0001). Chickens with limited microflora had higher gain:feed ratios (P = 0.002), higher fecal lipid digestibility (P = 0.0001), more intestinal conjugated bile salts (P = 0.0001), less intestinal unconjugated bile salts (P = 0.0001), and their gastrointestinal compartments were smaller (P < 0.05) than those of conventional chickens. Addition of bile salts in diets increased the lipid digestibility (P = 0.0001) with a more pronounced effect in conventional birds (P = 0.0001) and in birds fed on guar gum diets (P = 0.002). Feeding the guar gum diets increased the intestinal supernatant viscosity (P = 0.0001) for both microflora status. Guar gum addition increased (P < 0.05) the lactic acid concentration in the small intestine of conventional chickens. Guar gum decreased the fecal lipid digestibility (P = 0.0001) and the intestinal conjugated bile salts (P = 0.0001) for both microflora status. However, the magnitude of lipid digestibility decrease due to guar gum was lower (P = 0.05) in birds with limited microflora than in conventional birds, and the negative effect of guar gum on intestinal conjugated bile salts was more pronounced (P = 0.02) in birds with limited microflora. Bile salt addition reduced the negative effect of guar gum on lipid digestibility (P = 0.02) for both microflora status. The mean lipid digestibilities were negatively correlated (P = 0.0001) with the ratio [Ln(viscosity)/total bile salt] measured in intestinal digesta. Except for gizzard, size of digestive compartments and villus heights increased (P < 0.05) with feed efficiency and digestibility decreased. In conclusion, the results provided evidences that the negative effect of guar gum on lipid digestibility was mainly mediated by its direct effect on intestinal bile salt concentration and efficiency. The small interaction between guar gum and microflora status effects on lipid digestibility had to be accounted for by the low basal level of intestinal bile salts in conventional chickens.

Animal Feed↗
PubMed2006

Comparison of physiological and in vitro porcine gastric fluid digestion.

BACKGROUND: In previous studies, the major peanut allergen Ara h 1 was digested in vitro using pepsin and porcine gastric fluid. The results suggested that in vivo gastric digestion of allergen protein can be modeled accurately by peptic hydrolysis in vitro. In the current investigation, studies were designed to follow the gastrointestinal (GI) digestion of peanut allergens under true physiological conditions. In vitro digestion with porcine gastric fluid was compared with actual physiological digestion of peanut allergens in the porcine digestive tract in vivo. METHODS: Analysis of physiologic digestion was performed in piglets administered a 20-gram bolus of peanut meal followed by periodic sampling and analysis of GI contents. The pH was monitored, and digesta were analyzed by SDS-PAGE and immunoblot analysis. RESULTS: Peanut meal initially neutralized stomach contents to a pH of approximately 7, which was subsequently acidified by HCl secretion within 30 min. Acidification to pH 2-4 resulted in active pepsin digestion of soluble protein in the stomach. Soluble intact protein/allergens were rapidly degraded to pepsin-resistant peptides in the stomach followed by hydrolysis of these fragments in the small intestine. Particulate material was evident in both the stomach and small intestine that could contribute to continued release of peanut allergens Ara h 1, 2 and 3. CONCLUSIONS: Porcine gastric digestion of peanut proteins resembles true physiological digestion only under optimal physiologic conditions. Soluble proteins are rapidly digested and insoluble material continues to release IgE-reactive proteins throughout the GI tract. GI digestion of food allergens can play a prominent role when assessing allergens within the context of a food matrix or meal and during the sensitization phase of IgE-mediated allergy.

Allergens↗
PubMed1998

Effects of species raw material source, ash content, and processing temperature on amino acid digestibility of animal by-product meals by cecectomized roosters and ileally cannulated dogs.

We conducted experiments to determine amino acid (AA) digestibility of nine animal by-product meals using precision-fed cecectomized roosters and ileally cannulated dogs. The products initially evaluated in roosters were meat and bone meals (MBM) containing 24 or 34% ash, poultry by-product meals (PBP) containing 7 or 16% ash, lamb meals (LM) containing 15 or 24% ash, a LM analog containing a mixture of LM and turkey meal, and two MBM processed at either a low or high temperature. The MBM and PBP differing in ash, low-ash LM, and low-temperature MBM then were incorporated into extruded dry dog foods and evaluated in cecectomized roosters and ileally cannulated dogs. True digestibility of total AA in roosters averaged 76% for the nine meals fed alone, with the low-temperature MBM being highest at 84% and the low-ash LM being lowest at 66% (P < .05). No consistent differences in rooster AA digestibility were observed between pairs of meals differing in ash content. Digestibilities of AA were higher in the low-temperature MBM than in the high-temperature MBM. Differences in rooster AA digestibility values among the six extruded dog foods containing selected animal meals were similar to those observed when the animal meals were fed alone. The ileally cannulated dog assay yielded results for AA digestibilities that were highly correlated (r = .87 to .92) with those of the rooster assay, whereby the high-ash MBM and low-temperature MBM foods had the highest mean AA digestibility at 82% and the low-ash LM food had the lowest mean AA digestibility at 62% (P < .05). Again, no consistent differences in AA digestibilities for dogs were observed between pairs of dog foods containing MBM or PBP differing in ash content. Results of this study indicated that processing temperature influenced AA digestibility of MBM, but species raw material source and ash content had no consistent effect on AA digestibility. Results also indicated that the precision-fed cecectomized rooster assay could be used to predict differences in AA digestibility among animal by-product meals for dogs.

Amino Acids↗
PubMed2001

The effect of texturized vegetable protein from soy on nutrient digestibility compared to beef in cannulated dogs.

Texturized vegetable protein from soy (TVP) is widely used in canned dog foods, but its nutritional value remains in doubt. This study compared apparent prececal and total intestinal digestibility when four canned diets containing reciprocal proportions of protein from TVP (0 to 57%) and from beef (100 to 43%) were fed to eight cannulated dogs. As dietary TVP increased, the following linear changes were observed (P < 0.05): prececal and total intestinal protein digestibility decreased slightly from 77 to 71% and 86 to 80%, respectively; prececal amino acid digestibility decreased in parallel with protein; prececal carbohydrate digestibility decreased markedly from 80% to 62% and carbohydrate disappearance in the large intestine increased from 8 to 22%; prececal sodium digestibility decreased markedly (from 37 to 4%); and prececal potassium digestibility decreased (from 93 to 85%). Total intestinal digestibility of sodium and potassium decreased little (from 97 to 95% and from 98 to 97%, respectively); fecal mass and water content increased markedly (from 98 to 174 g/d and from 61 to 72%, respectively); and fat digestibility was unaffected. Prececal phosphorus digestibility from two diets containing TVP was lower than that from the all-beef diet (13 and 17%, vs 26%, P < 0.05). In conclusion, TVP is a useful source of protein in canine canned diets because amino acids from TVP are almost as digestible as those from beef in the canine intestine. Nevertheless, soy carbohydrate is poorly digested and large amounts of TVP inhibit small intestinal electrolyte digestibility and increase fecal water content.

Animals↗
PubMed2002

The influence of soy oligosaccharides on apparent and true ileal amino acid digestibilities and fecal consistency in growing pigs.

Fourteen ileally cannulated pigs (BW = 35 +/- 2 kg) were randomly allotted to a replicated 7 x 7 Latin square design experiment to evaluate the influence of the soybean oligosaccharides (OS), raffinose and stachyose, on ileal nutrient digestibility and fecal consistency. Semipurified diets containing soy protein concentrate (SPC) or soybean meal (SBM) as the sole protein sources were fed. Soy solubles (SS), a by-product of SBM processing containing 3.5% raffinose and 11.5% stachyose, were used to increase dietary raffinose and stachyose concentrations. The seven dietary treatments were SPC, SPC + 9% SS, SBM, SBM + 9% SS, SBM + 18% SS, SBM + 24,000 U alpha-galactosidase enzyme preparation/kg diet, and a low-protein casein (LPC) diet used to calculate true digestibility. Diets, with the exception of the LPC diet, were formulated to contain 17% CP. All diets contained 0.5% chromic oxide as a marker for ileal digestibility determination. The experimental periods were divided into a 5-d diet adaptation followed by 2-d of ileal digesta collection. Diets and digesta were analyzed for DM, N, Cr, amino acids (AA), raffinose, and stachyose. Fecal consistency was determined on d 6 and 7 of each experimental period. The apparent and true ileal AA digestibilities were not different (P < 0.05) for the SPC and SBM control diets. When SS was added to the SPC diet, apparent and true N and AA digestibilities were depressed (P < 0.05) with the exception of Trp and Pro. The apparent and true ileal N and AA digestibilities were not different (P > 0.05) between the SBM control and SBM + 9% SS diets with the exception of Glu. There was a linear decrease (P < 0.05) in apparent and true DM, Val, Gly, and Tyr digestibilities when increasing levels of SS were added to the SBM diet. The addition of alpha-galactosidase did not improve apparent or true ileal N or AA digestibilities except for apparent and true Val and Tyr. Ileal raffinose digestibility was improved (P < 0.05) by addition of a-galactosidase, but was not affected by any other dietary treatment. Ileal stachyose digestibility was not affected (P > 0.58) by treatment. Fecal consistency likewise was not affected (P > 0.36) by dietary treatment. In conclusion, soy OS reduced nutrient digestibilities, but the reductions were small, ranging from approximately 1.1 to 7.4 percentage units. This suggests that other factors may be negatively impacting SBM digestibility.

Amino Acids↗
PubMed1980

Digestibility of untreated and hydroxide-treated annual ryegrass straw.

Digestion trials with sheep were carried out with untreated and NaOH-treated cubed annual ryegrass straw. In the first trial, NaOH treatment increased (P < .05) digestibility of all organic components. Organic matter digestibility was increased by 21% and energy by 32%. In the second trial, the untreated straw (different crop year) was more digestible than in the first trial (digestible energy was 47%, vs 43% in the first year). In both cases, digestibility of the straw was estimated by difference when it was fed with alfalfa hay and compared to alfalfa hay alone. When the straw was fed with urea and molasses and digestibility was estimated with book values for molasses, digestion coefficients for organic matter and acid detergent fiber were slightly higher and energy was slightly lower than when digestibility was determined by difference, but the values were not different (P > .05). NaOH treatment did not result in as much improvement in the second trial as in the first trial, but it did result in increased digestibility or organic matter and fiber; crude protein digestibility however, was lower (P < .05) with NaOH-treated than with untreated straw. TDN estimated from digestible energy appeared to give more appropriate values for straw (39.7, 41.3) and NaOH-treated straw (44.4, 46.3) relative to alfalfa hay (53.4) than those calculated by the standard method (48.2 and 50.8 for straw; 52.0 and 55.0 for NaOH-treated straw).

Animal Feed↗
PubMed1986

Limits to starch digestion in the ruminant small intestine.

Site and extent of starch digestion by ruminant animals varies with species, grain type and processing method. Based on a review of 40 different experiments with cattle, between 18 and 42% of the dietary starch from corn and sorghum grains fed to cattle reaches the small intestine for digestion. With more extensive grain processing, a smaller quantity of starch reaches the small intestine. In the small intestine, from 47 to 88% of the presented starch is digested, while in the large intestine, 33 to 62% of the presented starch is digested. Though limits to digestion in and absorption from the small intestine can be demonstrated by infusing starch and glucose into the duodenum, enzymatic capacity does not appear to limit intestinal starch digestion since no plateau in the amount of starch disappearing from the small intestine is detected with typical diets. Yet, extent of digestion is incomplete. Other factors, such as time and surface exposure may limit small intestinal digestion of starch. Processing methods to reduce particle size or alter the protein matrix, which cements starch granules together, will increase the extent of digestion both in the rumen and in the small intestine. Performance data from growing cattle fed processed corn and sorghum grains indicate that starch was used 42% more efficiently if it was digested in the small intestine rather than in the rumen. Though total tract starch digestibility is of primary concern, results support the concept that energetic efficiency of growing ruminants is greater if starch is digested in the small intestine rather than in the rumen.

Animal Feed↗
PubMed1997

Effects of feeding nonforage fiber sources on site of fiber digestion.

Although many nonforage fiber sources have high extents of neutral detergent fiber (NDF) digestion, most have rates of digestion similar to or slower than the rates of forage NDF digestion. Rates of NDF digestion vary considerably among and within sources of by-products. Digestion kinetics also vary because of the technique used (in vitro versus in situ) and because of high amounts of dietary concentrate. Based on available data for passage rate and specific gravity measurements, rates of passage of nonforage fiber sources from the rumen of high producing cows appear to be faster than those of forages. Therefore, the potential to shift NDF digestion to the hindgut has been discussed. To account for variability in ruminal and total tract digestibility of NDF, multiple regression analysis was used to indicate that nonforage NDF percentage in the diet had about two-thirds the positive response on total tract NDF digestion that forage NDF percentage did. Although the loss of potentially digestible NDF may occur, DMI does not appear to decrease much until forage NDF is below 14 to 16% of dietary DM. Conversely, replacement of starch with nonforage NDF appears to increase digestibility of fiber, mostly in diets with high concentrations of nonfiber carbohydrates, apparently because of reduced negative associative effects. Increasing the concentration of total NDF above 35% also can decrease DMI with little improvement in NDF digestibility. Increased knowledge of the kinetics of digestion and the passage of various nonforage fiber sources used to replace forage or concentrate should increase the accuracy and precision of dynamic models, thereby increasing the flexibility and utility of nonforage fiber sources in dairy rations.

Animals↗
PubMed1992

[The effect of copper and cobalt supplementation on the digestibility of fibrous feed in sheep].

The effects of an addition of copper, cobalt, and copper + cobalt on the in vitro digestibility of hay, treated beech sawdust and cellulose by the rumen fluid of sheep fed a complete food ration, complete food ration with 15% treated beech sawdust and a diet consisting of 80% hay and 20% barley were investigated, respectively. The rumen fluid obtained from animals fed a complete food ration showed the significantly higher digestibility of hay (Tab. I) when Cu (P less than 0.05), Co, or Cu + Co (P less than 0.001) had been added into the rumen. The cellulose digestibility increased only after Co application (P less than 0.05). The microelements supplementation in all three forms had a very significant effect on the digestibility of treated beech sawdust by the rumen fluid of sheep fed a diet with 15% sawdust content, although the initial digestibility by the rumen fluid in the control animals (without microelements) was only 44.9%. A similar trend was also observed in the cellulose digestibility, but after Co application this increase was not significant. The rumen fluid of animals fed hay (80%) and barley (20%) showed the highest digestibility. The digestibility of sawdust reached 60% and the of cellulose 95.9% without microelements supplementation (control). In this case the effect of microelement supplementation on the sawdust digestibility was manifested only in the case of cobalt by the digestibility decrease (P less than 0.001). The cellulose digestibility also decreased after administration of Cu (P less than 0.01) and Co (P less than 0.05). The Cu + Co supplementation did not influence its digestibility.(ABSTRACT TRUNCATED AT 250 WORDS)

Animal Feed↗
PubMed1980

[Comparative study of the digestion of a complete food in the pony and the rabbit].

A complete pelleted diet (table 1) containing 11 p. 100 of crude protein and 17 p. 100 of cellulose (ADF-lignine according to the method of Van Soest) was distributed to ponies and rabbits. Total digestibilities, partial cumulative digestibilities in different compartments of the digestive tract (stomach, small intestine, caecum, proximal and distal colon) and changes in biochemical composition, measured by pH, VFA, lactic acid and ammonia concentrations, were compared. Total digestibilities (table 2). There was no significant difference in dry matter between the two species. Crude protein digestibility was higher in ponies, but a greater difference was observed for crude fiber digestibility: digestibility in the rabbit was 60 p. 100 of that in the pony. Starch was entirely digested in ponies and rabbits. Partial digestibilities (table 3). The dry matter content was the same in the stomach, the different parts of the small intestine and the caecum of both species. Starch was rapidly digested, but crude protein digestibility was highly negative in the stomach, small intestine and caecum of both species. Changes in biochemical composition (table 4). VFA concentrations were higher in the caecum and colon of ponies but molar percentages of acetic, propionic and butyric acids were very comparable (fig. 1); L-lactic acid concentration was higher in the first part of the digestive tract of ponies and ammonia concentration was lower.

Ammonia↗
PubMed1986

Effects of diet concentrate level and sodium bicarbonate on site and extent of forage fiber digestion in the gastrointestinal tract of wethers.

Four adult wethers (45 kg) with permanent ruminal and abomasal cannulae were used in a repeated measures Latin-square arrangement of treatments to quantitate the effects of diet concentrate level and sodium bicarbonate (NaHCO3) on site and extent of forage fiber digestion in the gastrointestinal tract. Experimental diets consisted of Kentucky-31 tall fescue hay, soybean meal and a semi-purified concentrate mixture in ratios of 95:5:0, 76:4:20, 57:3:40 and 38:2:60; NaHCO3 represented 0 or 7.5% of the concentrate mixture. Ruminal digestion (% of intake) of neutral detergent fiber (NDF) and hemicellulose decreased linearly (P less than .05), whereas acid detergent fiber (ADF) digestion responded in a cubic (P less than .05) fashion to increasing concentrate level; NaHCO3 improved ruminal digestion of NDF (P less than .10) and ADF (P less than .05), but not hemicellulose. Post-ruminal digestion (% of rumen non-degraded) of NDF and ADF tended to increase, whereas hemicellulose digestion responded in a cubic (P less than .05) fashion to increasing concentrate level; NaHCO3 decreased (P less than .05) post-ruminal digestion of all fiber fractions. Total tract digestion of NDF and ADF showed a cubic (P less than .05) response, whereas hemicellulose digestion responded in a quadratic (P less than .05) fashion to increasing concentrate level; NaHCO3 had no effect on total tract digestion of any fiber fraction. Correlations of ruminal hemicellulose digestion with mean pH (r = .33; P = .07) and minimum pH (r = .30; P = .09) were attained in a 24-h feeding cycle.(ABSTRACT TRUNCATED AT 250 WORDS)

Animal Feed↗
PubMed2002

Steam pressure disruption of municipal solid waste enhances anaerobic digestion kinetics and biogas yield.

Biomass waste, including municipal solid waste (MSW), contains lignocellulosic-containing fiber components that are not readily available as substrates for anaerobic digestion due to the physical shielding of cellulose imparted by the nondigestible lignin. Consequently, a substantial portion of the potentially available carbon is not converted to methane and the incompletely digested residues from anaerobic digestion generally require additional processing prior to their return to the environment. We investigated and developed steam pressure disruption as a treatment step to render lignocellulosic-rich biomass more digestible and as a means for increasing methane energy recovery. The rapid depressurization after steam heating (240 degrees C, 5 min.) of the nondigested residues following a 30-day primary digestion of MSW caused a visible disruption of fibers and release of soluble organic components. The disrupted material, after reinoculation, provided a rapid burst in methane production at rates double those observed in the initial digestion. This secondary digestion proceeded without a lag phase in gas production, provided approximately 40% additional methane yields, and was accompanied by a approximately 40% increase in volatile solids reduction. The secondary digestate was found to be enriched in lignin and significantly depleted in cellulose and hemi-cellulose components when compared to primary digestate. Thus, steam pressure disruption treatment rendered lignocellulosic substrates readily accessible to anaerobic digestion bacteria and improved both the kinetics of biogas production and the overall methane yield from MSW. Steam pressure disruption is central to a new anaerobic digestion process approach including sequential digestion stages and integrated energy recovery, to improve process yields, provide cogenerated energy for process needs, and to provide effective reuse and recycling of waste biomass materials.

Anaerobiosis↗
PubMed1988

Effects of alpha-galactosidase digestion on lectin staining in human pancreas.

Effects of alpha-galactosidase (from green coffee beans) digestion on lectin staining were examined in formalin-fixed, paraffin-embedded human pancreatic tissues from individuals of blood-group B and AB. Digestion with the enzyme resulted in almost complete loss of Griffonia simplicifolia agglutinin I-B4 (GSAI-B4) staining in the acinar cells with concomitant appearance of Ulex europaeus agglutinin-I(UEA-I) staining in the corresponding cells. In addition, reactivity with soybean agglutinin(SBA) was also imparted by the enzyme digestion in GSAI-B4 positive acinar cells. beta-Galactosidase digestion following alpha-galactosidase digestion neither reduced the reactivity with SBA nor induced the reactivity with Griffonia simplicifolia agglutinin-II(GSA-II) in GSAI-B4 positive cells, while in UEA-I positive cells, both reduction of SBA reactivity and appearance of GSA-II reactivity occurred after simple beta-galactosidase digestion as well as sequential digestion with alpha- and beta-galactosidase. However, when alpha-L-fucosidase digestion procedure was inserted between alpha- and beta-galactosidase digestion, UEA-I staining imparted by alpha-galactosidase digestion was markedly decreased in intensity and GSA-II reactivity was appeared in GSAI-B4 positive acinar cells. Furthermore, after sequential digestion with alpha-galactosidase and fucosidase, reactivity with peanut agglutinin(PNA) was revealed in GSAI-B4 positive acinar cells as well as UEA-I positive cells in secretors. In non-secretors, strong PNA staining was usually observed in the acinar cells throughout the glands without enzyme digestion.(ABSTRACT TRUNCATED AT 250 WORDS)

ABO Blood-Group System↗
PubMed2006

A bifunctional monolithic column for combined protein preconcentration and digestion for high throughput proteomics research.

Enzymatic digestion of proteins is a key step in protein identification by mass spectrometry (MS). Traditional solution-based protein digestion methods require long incubation times and are limitations for high throughput proteomics research. Recently, solid phase digestion (e.g. trypsin immobilization on solid supports) has become a useful strategy to accelerate the speed of protein digestion and eliminate autodigestion by immobilizing and isolating the enzyme moieties on solid supports. Monolithic media is an attractive support for immobilization of enzymes due to its unique properties that include fast mass transfer, stability in most solvents, and versatility of functional groups on the surfaces of monoliths. We prepared immobilized trypsin monolithic capillaries for on-column protein digestion, analyzed the digested peptides through LC/FTICR tandem MS, and compared peptide mass fingerprinting by MALDI-TOF-MS. To further improve the digestion efficiency for low abundance proteins, we introduced C4 functional groups onto the monolith surfaces to combine on-column protein enrichment and digestion. Compared with immobilized trypsin monolithic capillaries without C4, the immobilized trypsin-C4 monolith showed improved digestion efficiency. A mechanism for increased efficiency from the combination of sample enrichment and on-column digestion is also proposed in this paper. Moreover, we investigated the effects of organic solvent on digestion and detection by comparing the observed digested peptide sequences. Our data demonstrated that all columns showed good tolerance to organic solvents and maintained reproducible enzymatic activity for at least 30 days.

Animals↗
PubMed2003

Mass spectrometric study of the effects of hydrophobic surface chemistry and morphology on the digestion of surface-bound proteins.

Our previous work has demonstrated that reversed-phase chromatographic micro-beads can be used to capture proteins from complex biological matrices and the surface-bound proteins can be enzymatically digested for protein identification by mass spectrometry (MS). Here we examine the peptides generated from digestion of proteins bound to various types of micro-bead surfaces in order to determine the effects of surface chemistry and surface morphology on the digestion process. Detailed examinations of site cleavages and sequence coverage are carried out for a tryptic digestion of cytochrome c adsorbed on reversed-phase polystyrene divinylbenzene (Poros R2 beads) versus C(18) bonded-phase silica beads. It is shown that although the surface does not completely hinder the digestion of cleavage sites of the protein, the digestion products are clearly different than those obtained from a solution digest. Specifically, a partial digestion results from surface digestion, resulting in a greater number of missed cleavages than a comparable solution digest. Subsequent comparisons of peptide mass maps generated from the digestion of various proteins on surfaces with altering chemistry (C(4), C(8), C(18), and R2 beads), or with different surface morphology, were performed. The results reveal that surface chemistry plays only a minor role in affecting the peptide mass maps, and surface morphology had no noticeable effects on the resulting peptide mass maps. It is also shown that the mass spectrometric detection method used to analyze the digested peptides can significantly influence the information content on cleavage sites and the extent of sequence coverage. The use of a combination of MALDI, LC/off-line MALDI, and LC/ESI MS is demonstrated to be crucial in revealing subtle changes in the peptide mass maps.

Chromatography, High Pressure Liquid↗
PubMed2006

Efficient and specific trypsin digestion of microgram to nanogram quantities of proteins in organic-aqueous solvent systems.

Mass spectrometry-based identification of the components of multiprotein complexes often involves solution-phase proteolytic digestion of the complex. The affinity purification of individual protein complexes often yields nanogram to low-microgram amounts of protein, which poses several challenges for enzymatic digestion and protein identification. We tested different solvent systems to optimize trypsin digestions of samples containing limited amounts of protein for subsequent analysis by LC-MS-MS. Data collected from digestion of 10-, 2-, 1-, and 0.2-microg portions of a protein standard mixture indicated that an organic-aqueous solvent system containing 80% acetonitrile consistently provided the most complete digestion, producing more peptide identifications than the other solvent systems tested. For example, a 1-h digestion in 80% acetonitrile yielded over 52% more peptides than the overnight digestion of 1 microg of a protein mixture in purely aqueous buffer. This trend was also observed for peptides from digested ribosomal proteins isolated from Rhodopseudomonas palustris. In addition to improved digestion efficiency, the shorter digestion times possible with the organic solvent also improved trypsin specificity, resulting in smaller numbers of semitryptic peptides than an overnight digestion protocol using an aqueous solvent. The technique was also demonstrated for an affinity-isolated protein complex, GroEL. To our knowledge, this report is the first using mass spectrometry data to show a linkage between digestion solvent and trypsin specificity.

Acetonitriles↗