Pancreatin Digestion Research Articles

Binding of iron by chicken muscle protein digests: the size of the iron-binding peptides

The dilute salt-insoluble proteins (DSIPs) were extracted from chicken muscle and digested in the presence of ferric iron. The size of the peptides was determined by ultrafiltration using membranes with various molecular weight cut-offs (MWCOs). Over 90% of the peptides in both the pepsin and pepsin–pancreatin digestions were smaller than 10 kDa and more than 45% were smaller than 3.5 kDa. Pancreatin digestion increased the amount of small peptides (<3.5 kDa) as compared to pepsin digestion alone. Most of the iron was in a form too large to pass the 10 kDa MWCO membrane and was presumably bound to a few large peptides. About 10% of the iron was smaller than 3.5 kDa, but there was very little in the intermediate range (3.5–10 kDa). Pancreatin digestion produced more soluble iron than pepsin alone, but the size distribution was similar for both digestions. The smaller peptides (<10 kD) bound very little iron during digestion but could bind iron if separated from the large peptides before iron addition. The large iron-binding peptides were separated by ultrafiltration, and a 22-fold increase in specific activity (iron mg−1 peptide) was achieved using a 10 kDa MWCO membrane. © 2000 Society of Chemical Industry

Phosphorus bioavailability and digestibility of normal and genetically modified low-phytate corn for pigs.

We conducted two studies to determine the bioavailability and apparent digestibility of P in a low-phytate corn hybrid (.28% total P, .10% phytate P) genetically modified to be homozygous for the 1pa1-1 allele and a nearly isogenic corn hybrid (normal) (.25% total P, .20% phytate P). Additionally, we conducted an in vitro assay involving a peptic and pancreatin digestion to estimate P availability. The first study used 50 individually penned pigs (initial body weight 9 kg) and 10 treatments in a randomized complete block design. A cornstarch-soybean meal basal diet (.6% Ca, .2% P) was used. Treatments consisted of the basal diet and the basal diet plus .05, .10, or .15% P from monosodium phosphate (MSP), low-phytate corn, or normal corn. After a 35-d feeding period, pigs were killed to collect the fourth metacarpal for measurements of ash and breaking load. Breaking load was regressed on added P intake, and the bioavailability of P was determined by the slope ratio method. The bioavailabilities of P (relative to MSP) for low-phytate and normal corn were 62 and 9%, respectively. These were similar to the determined in vitro values of 57 and 11% for low-phytate and normal corn, respectively. In the second study, 20 pigs (initial BW 20 kg) were used in a randomized complete block design with a 2 x 2 factorial arrangement of treatments. Two corn lines (low-phytate and normal) and two levels of supplemental P (0 and .2%) from dicalcium phosphate were used. Diets with no added P were formulated to contain .9% lysine, .6% Ca, and .34% P. Apparent nutrient digestibilities were calculated from total collection of urine and feces for 5 d. There were no differences among treatments for energy and nitrogen digestibility. Pigs fed low-phytate corn with no added P had increased digestibility and retention of P and reduced total P excretion (P < .05). We conclude that low-phytate corn contains at least five times as much available P as normal corn. The use oflow-phytate corn greatly reduced the amount of P excreted by the pig and increased the N:P ratio in the manure.

Study on effect and mechanism of essential garlic oil in inhibiting monocyte-vascular endothelial cell adhesion induced by interleukin-1α

To observe the effects of essential garlic oil (EGO) on vascular cell adhesive molecule-1 (VCAM-1) expression of endothelial cell and monocyte adhesion induced by interleukin-1 alpha (IL-1 alpha). Human umbilical vein endothelial cells (HUVEC) were isolated by pancreatin digestion method and co-cultured with U937 monocyte and IL-1 alpha or EGO + IL-1 alpha respectively. The adhesion rate of monocyte to endothelial cell was measured and mean fluorescent intensity of immuno-stained of VCAM-1 was determined by ACAS 570. EGO showed significant inhibition on VCAM-1 expression induced by IL-1 alpha either in simple culture with HUVEC or in co-culture with HUVEC + U937, and could obviously down regulate the monocyte-endothelial cell adhesion under effect of IL-1 alpha. EGO has the effect of antagonizing against adhesion of monocyte and vascular endothelial cell, it may be due to the inhibition of EGO on adhesive molecular expression on endothelial cell.

Procedures for the isolation and culture of Sertoli cells from the testes of infant, juvenile, and adult rhesus monkeys (Macaca mulatta).

The purpose of the present study was to establish culture conditions for the in vitro study of the rhesus monkey Sertoli cell (Sc) at three major stages of development, namely infancy, adulthood, and the intervening prepubertal period. Conditions for the culture of Sc from juveniles were first established using collagenase and pancreatin digestion of seminiferous tubules. The addition of 1% fetal bovine serum for the first 24 h of culture was necessary for attachment of Sc clusters. Confluency of Sc from juveniles was reached as early as 4 days of culture. Histochemical and ultrastructural observations confirmed that the cultures were enriched with Sc and that contamination by peritubular cells was minimal (2%). Although application of similar culture conditions was successful in establishing cultures of Sc from infants, significant modification of the procedure was required before Sc from adults could be cultured. Specifically, adult testicular tissue required two sequential collagenase digestions at elevated temperature. The yield of adult Sc, however, remained low. Cultures of juvenile Sc produced substantial quantities of 31-kDa inhibin, which was bioactive as reflected by its ability to suppress FSH secretion from rat pituitary cells in vitro. Although aromatase activity in juvenile Sc cultures was stimulated by FSH, inhibin synthesis, as reflected by immunoactive inhibin production and steady-state levels of alpha inhibin mRNA, was not increased by FSH. The establishment of conditions for the culture of infant, juvenile, and adult Sc from the rhesus monkey will provide a model for study of the postnatal ontogeny of Sc function in higher primates.

Effect of retrogradation, pancreatin digestion and amylose/amylopectin ratio on the fermentation of starch byClostridium butyricum (NCIMB 7423)

Using three different maize starches (maize, waxy maize and high amylose maize, containing 25%, 1% and 52% amylose, respectively) the influence of amylose/amylopectin content and of retrogradation on fermentation by the porcine caecal anaerobe Clostridium butyricum was assessed. Small intestine digestion was simulated using pancreatin before the starches were exposed to bacterial fermentation. It was found that retrogradation appeared to alter the extent of the fermentation and hence the amount of short-chain fatty acids produced, while pancreatin digestion appeared to alter the way in which the organism fermented the starch and hence the acetate/butyrate ratio. The amylose/amylopectin ratio seemed to have more influence on the way the starch was fermented by the bacteria after the starch had been subjected to digestion with pancreatic enzymes, but had less influence when the starch had been retrograded. © 1998 SCI.

Effect of retrogradation, pancreatin digestion and amylose/amylopectin ratio on the fermentation of starch by Clostridium butyricum (NCIMB 7423)

Effect of retrogradation, pancreatin digestion and amylose/amylopectin ratio on the fermentation of starch by Clostridium butyricum (NCIMB 7423)

Effect of calcium and zinc concentrations and calcium source on in vitro calcium and zinc solubility in a fiber-fortified enteral formula

The present study was carried out to evaluate the effects of simultaneous addition of increasing levels of calcium (60–210 mg/100 g formula) and zinc (1.5–3.9 mg/100 g formula) on calcium and zinc solubility in an enteral formula fortified with soy polysaccharide fiber (2.13 g/100 g formula). The solubility of calcium from various sources, added to the formula, was also examined. Solubility was estimated by the ratio of soluble calcium or zinc in the supernatant, after centrifugal separation of samples subjected to in vitro pepsin and pancreatin digestion, over the total calcium or zinc content of samples. No interaction was found between calcium and zinc levels. Zinc solubility decreased linearly from 17 to 12% with increasing calcium levels (p<0.05). Soluble calcium decreased from 18 to 13% with increasing zinc levels (p<0.01). The percentage of soluble zinc was lower as more zinc was added (p<0.01) but soluble calcium was higher as more calcium was added (p<0.01). Mono- and dibasic calcium phosphates were markedly less soluble than calcium carbonate, calcium chloride and calcium phosphate hydroxide. Results suggest a competitive effect between calcium and zinc when soy polysaccharide fiber is present. An understanding of this effect is necessary for the appropriate fortification of enteral formulas in order to maximize the proportion of minerals potentially available for absorption.

Identification of an Antihypertensive Peptide from Casein Hydrolysate Produced by a Proteinase from Lactobacillus helveticus CP790

Casein hydrolysate, produced by an extracellular proteinase from Lactobacillus helveticus CP790, was fractionated by two-step reverse-phase HPLC. Only one fraction showed antihypertensive activity as measured by systolic blood pressure in spontaneously hypertensive rats after oral administration. Ten peptides in the fraction were further purified and identified by analysis of amino acid sequences. Each identified peptide was chemically synthesized, and the antihypertensive activity of each peptide was evaluated in spontaneously hypertensive rats. The synthetic peptide with a sequence of Lys-Val-Leu-Pro-Val-Pro-Gln, found in β-casein, indicated strong anti-hypertensive activity from 2 to 10h after oral administration of 2mg of peptide/kg of BW, and the effect was maximal at 6h after oral administration (−31.5±5.6mm Hg). Moreover, the antihypertensive effect of the peptide was dependent on the dosage of peptide from 0.5 to 2mg of peptide/kg of BW. Interestingly, the antihypertensive peptide showed lower inhibitory activity of angiotensin I-converting enzyme, but the activity was increased after pancreatin digestion.

Fermentation of native and processed starches by the porcine caecal anaerobe Clostridium butyricum (NCIMB 7423)

Starch fermentation by the porcine caecal anaerobe Clostridium butyricum was examined using gas and volatile fatty acid production as determinants of activity. Potato starch and amylopectin were studied in their native form as well as after retrogradation, which should render them resistant to pancreatic α‐amylase digestion. Fermentation of both substrates was enhanced by pancreatin digestion of the native material, possibly due to the removal or disruption of part of the structure of the starch by the pancreatic enzymes. However, pancreatic digestion of retrograded potato starch apparently reduced the amount available for bacterial fermentation, whereas no significant effect was observed with amylopectin. The data suggests that starches which are high in amylopectin would be more likely to influence fermentation in the large intestine in monogastric animals, and that the presence of residual pancreatic enzymes in the lower gut could potentially enhance starch fermentation by this micro‐organism.

An in vitro procedure for studying enzymic dephosphorylation of phytate in maize-soyabean feeds for turkey poults

An in vitro method was developed to predict inorganic P release from maize-soyabean poultry feeds containing supplemental phytase (EC 3.1.3.8), and to quantify the effect of acid phosphatase (EC 3.1.3.2), fungal protease (EC 3.4.23.6) and Aspergillus niger cellulase (EC 3.2.1.4) on phytate dephosphorylation. Pepsin (EC 3.4.23.1) and pancreatin digestion periods were preceded by a 30 min pre-incubation at pH 5.25 to simulate digestion in the crop of poultry. Pancreatin digestion was carried out in dialysis tubing, with a ratio of about 1:25 (v/v) between the digesta and dialysing medium, to simulate gradient absorption from the duodenum. The feed:water ratio was kept within physiological limits and a constant proportion of feed weight to digestive enzymes was maintained. There was a linear response to increasing dosages of phytase up to 1000 phytase units (FTU)/kg feed, and to increasing phosphate concentration in feeds. In vivo validation was performed with growing turkeys (1-3 weeks) fed on diets containing 12 g Ca/kg and 0, 500 or 1000 FTU phytase/kg in a factorial arrangement with 0, 1, 2 or 3 g supplemental phosphate/kg (from KH2PO4). After a simple transformation (variable/in vitro P = f (in vitro P)), amounts of P hydrolysed from feed samples by in vitro digestions correlated with 3-week body-weight gain (R 0.986, P < 0.0001), toe ash (R 0.952, P < 0.0001), feed intake (R 0.994, P < 0.0001) and feed efficiency (R 0.992, P < 0.0001). The dephosphorylating ability of phytase in vitro was significantly enhanced (P < 0.05) by the addition of acid phosphatase. Fungal acid protease and Aspergillus niger cellulase also enhanced the dephosphorylation process in vitro.

A three-step in vitro procedure for estimating intestinal digestion of protein in ruminants2

A three-step in vitro procedure was developed to estimate intestinal digestion of proteins in ruminants. Dacron bags containing feed samples were suspended in the rumen for 16 h. Residue containing 15 mg of N after ruminal exposure was incubated for 1 h in 10 mL of a .1 N HCl solution containing 1 g/L of pepsin. After incubation, pH was neutralized with .5 mL of 1 N NaOH and 13.5 mL of a pH 7.8 phosphate buffer containing 37.5 mg of pancreatin were added to the solution and incubated at 38 degrees C. After a 24-h incubation, 3 mL of a 100% (wt/vol) trichloroacetic acid solution were added to precipitate undigested proteins. Preincubation of samples in the rumen did not affect (P > .05) pepsin-pancreatin digestion of residual CP in soybean meal (SBM), corn gluten meal (CGM), and blood meal (BM) and reduced (P < .05) pepsin-pancreatin digestion of residual CP in hydrolyzed feather meal (HFM), fish meal (FM), and meat and bone meal (MBM) (80 vs 70, 88 vs 81, and 82 vs 56%, respectively, for nonruminal vs ruminal preincubation). Pepsin digestion before pancreatin digestion increased (P < .05) CP digestion of all proteins tested by a mean of 23 percentage units. The pancreatin digestion step was validated using 34 duodenal samples from which small intestinal CP digestion was determined in vivo. The regression equation of in vivo estimates on pancreatin digestion had an r value of .91 (P < .001).(ABSTRACT TRUNCATED AT 250 WORDS)

METHIONINE LIBERATION BY PEPSIN-PANCREATIN HYDROLYSIS OF BEAN PROTEIN FRACTIONS: ESTIMATION OF METHIONINE BIOAVAILABILITY

Albumin (ALB), globulin (GLO), globulin G1 (G1), glutelin (GLU), and a protease inhibitor-lectin rich fraction (PIL) were extracted from IAC-Carioca 80 SH (Phaseolus vulgaris) based on differential solubility. The purpose of the study was to search for different susceptibilities to proteolysis and methionine liberation among protein fractions. The fractions were submitted to pepsin digestion for 3 h, then to pancreatin digestion for another 3 h, under optimum pH and 37C. Degree of hydrolysis (DH) was followed by the TNBS reaction and methionine liberation (Met%) by the chloramine-T method. DH for undenatured proteins was higher for GLO, Gl and GLU (62–71%) and lower for ALB and PIL (51–53%). After denaturation, DH was higher for Gl, GLU and GLO (81–86%) and lower for PIL and ALB (66–78%). For undenatured proteins Met% was higher for GLO, Gl and GLU (42–48%), and lower for ALB and PIL (22–29%). For denatured proteins Met% was higher for Gl, ALB and GLO (66–73%) and lower for PIL and GLU (50–53%). Overall, hydrolysis and methionine liberation followed the same general pattern for both pepsin and pancreatin digestion, individually.

Assessment of soluble and ionic calcium released from milk and dairy products during simulated gastro-intestinal digestion

Abstract The amounts of soluble and ionic calcium released from dairy products have been measured during simulated gastro-intestinal digestion. Different dairy products, including skim milk, evaporated milk, yogurt and cheese, containing various concentrations and forms of calcium, were submitted to pepsin and pancreatin digestion using a dialysis cell unit. Digestion products were continuously collected for quantitative determination of soluble and ionized calcium released. The results obtained showed the importance of pepsin digestion on the potential availability of dietary calcium from dairy products. Moreover, data have indicated that lower amounts of soluble and ionized calcium were released from cheese than from yogurt and evaporated milk during simulated gastro-intestinal digestion.

VERIFICAÇÃO DAS CONDIÇÕES HIGIÊNICAS DE PRODUTOS DE CONFEITARIA*

Foram analisadas 60 amostras de 4 tipos diferentes de produtos de confeitaria: panetones, pães doces recheados ou não, bolos e roscas adquiridas em supermercados e padarias da Capital, são Paulo, com a finalidade de se verificar a sanidade destes produtos. Nesta pesquisa ressaltou-se a importância do desenvolvimento e de adaptações de métodos às condições de trabalho. Os resultados obtidos mostraram que 27 (45,0%) das amostras estavam em condições higiênicas in satisfatórias ou impróprias para consumo, sendo que 15 (25,0%) continham mais de 30 fragmentos de insetos, 9(15,0%) continham pêlos de roedor, 6 (10,0%) estavam contaminadas com insetos, 5 (8,3%) com larvas e 5 (8,3%) continham ácaros. Concluiu-se que os produtos industrializados estavam em melhores condições higiênicas do que os produtos de confeitaria.

A new rapid enzyme digestion method for predicting in Vitro protein quality (PDD index)

The officially accepted Protein Efficiency Ratio (PER) assay has limited applicability as a quality control and process monitoring assay due to the 28 days required to complete the assay. The ultimate measure of protein nutritional quality is the human bio-assay; however, this takes from 35–45 days to complete and is very expensive compared to the PER assay. PER is recognised by many nutritional biochemists as only an interim measure of human protein quality. The need for a reliable, timely, inexpensive rapid protein quality assay has become more evident in recent years with many rapid methods, including the Pepsin Digest Residue (PDR), Pepsin Pancreatin Digest (PPD), Pepsin Pancreatin Digest Dialysate (PPDD) and the Calculated Protein Efficiency Ratio (C-PER) methods, appearing in the research literature over the past 20 years. While each assay has notable merits, inherent limitations associated with each prevent use on an industrial basis. We now report a rapid assay, the Pepsin Digest Dialysate (PDD) Index, that can be used to determine protein quality over a wide range of protein sources currently available in human foods and which is acceptable for use in the food and nutritional product industries and for the Food and Drug Administration.

Effect of Enzymatic Digestion, pH and Molecular Weight on the Iron Solubilizing Properties of Chicken Muscle

ABSTRACTThe effects of a simulated gastrointestinal pH, enzymatic digestion and molecular weight (MW) on the iron solubilizing properties of a heated dilute salt insoluble fraction of chicken muscle were examined. The solubility of 50 ppm added FeCl3 increased linearly from 0–260 min during pepsin digestion. The total soluble iron reached a maximum concentration following a 120 min pepsin‐30 min pancreatin digestion, with pepsin digestion products ranging in MW from 6200–2500. Solubilization capacity, defined as an in vitro measure of total iron bioavailability, did not correlate to binding by free sulfhydryl groups. The soluble low molecular weight iron chelates found may explain, in part, the mechanism by which the “meat factor” enhances iron bioavailability.

Solubility of inorganic iron as affected by proteolytic digestion

The effects of enzymatic digestion on the iron-solubilizing properties of chicken muscle were examined. A water-soluble extract, an acid-soluble extract, and an acid-insoluble fraction were subjected to a simulated gastrointestinal digestion using pancreatin and/or pepsin: The solubility of added Fe was significantly affected only by the acid-insoluble fraction and increased linearly as pepsin digestion progressed from 0 to 4 h. A maximum was reached when this treatment was followed by a 1-h pancreatin digestion. Pepsin digestion products with molecular weight (MW) less than 10,000 solubilized significantly more Fe than those with MW greater than or equal to 10,000. In contrast pancreatin digestion products of MW less than 10,000 were not effective Fe-solubilizing agents. The influence of chicken breast muscle on added Fe solubility appears to be related to the production of digestion intermediates that can act as ligands in the formation of soluble Fe complexes.

Effects of Free and Bound Chlorogenic Acid on the In Vitro Digestibility of Ribulose Bisphosphate Carboxylase from Spinach

ABSTRACTThe effect of free and bound chlorogenic acid on the in vitro digestibility of spinach ribulose bisphosphate carboxylase from spinach or fraction 1 (F‐1) protein was investigated using a pH stat method. The pancreatin digestibility of F‐1 protein was less than that of casein and BSA but greater than 11s soy protein. Noncovalently bound chlorogenic acid increased the initial rate of hydrolysis using both pancreatin and trypsin. Viscosity measurements suggested non‐ covalent binding of chlorogenic acid caused partial unfolding of F‐1 protein making it more susceptible to proteolysis. Covalently bound chlorogenic acid decreased the rates of hydrolysis of F‐1 by both trypsin and pancreatin, and this was correlated with changes in available lysine.

Proteolytic activity, amino acid composition and protein quality of germinating fenugreek seeds ( Trigonella foenum graecum L.)

This study determined the changes in proteolytic activity and trypsin inhibitor, due to germination for 120 h, and their relationships to amino acid content and protein quality. The activities of cas-ase, Hb-ase, and BAPA-ase, as well as the trypsin inhibitor of the fenugreek, increased as germination proceeded. Glutamic and aspartic acids were the richest amino acids in fenugreek seeds, followed by leucine, arginine, lysine and proline. After germination, the contents of aspartic acid, phenylalanine, tyrosine, threonine, tryptophan and valine increased, while those of glutamic acid, proline and four of the essential amino acids decreased. All the free amino acids increased after germination, while glycine decreased. Cystine, tryptophan and proline originated in a free form after germination. The chemical score, based on the essential amino acid pattern of whole egg protein, for fenugreek seeds, showed a marked decrease (21%) after germination. The in vitro pepsin, followed by pancreatin digestion, showed a slight increase for the germinated seeds. Germination of seeds lowered the C-PER, a result which indicates that the protein quality decreased.

Biosynthesis and regulation of iodolipids in calf thyroid.

The incorporation of 125I into iodolipids was investigated in calf thyroid slices. Time course studies showed that the iodination of lipid reaches a plateau after 30 min of incubation. A highly significant correlation was found between iodination of lipid and of protein (r = 0.906), suggesting that both reactions may be related. Addition of PTU or MMI caused a significant inhibition of lipid iodination, indicating that a peroxidase could be involved in this reaction. The iodinated lipids are not attached to protein, since they migrated in BEA chromatography and pancreatin digestion of the samples did not modify the percentage radioactivity. The iodolipid fraction observed in BEA chromatography was eluted and analysed by TLC. Iodinated free fatty acids and neutral lipids comprised most of the radioactivity. Although iodolipids are present in every subcellular fraction, the 20 000 X g pellet had the greatest proportion of iodinated fatty acids and neutral lipids.