PREVIOUS work has shown that ingestion

Size: px

Start display at page:

Download "PREVIOUS work has shown that ingestion"

Carmella Gilmore
5 years ago
Views:

1 192 B. C. DILWORTH, C. D. SCHULTZ AND E. J. DAY Summit, Pennsylvania, for their cooperative efforts and grant-in-aid in support of this work. REFERENCES Dilworth, B. C, C. D. Schultz and E. J. Day, Salt utilization studies with poultry. 1. Effect of salt sources, particle size and insolubles in broiler performance. Poultry Sci. 49: Duncan, D. B., Multiple range and multiple F test. Biometrics, 11: Griffith, M., Effect of dietary particle size on phosphorus availability. Poultry Sci. 47: Snedecor, G. W., Statistical Methods, 5th Edition, Iowa State College Press, Ames, Iowa. Lipase in and Intestinal Contents of Chickens Heated and Raw Soybean s S. LEPKOVSKY AND F. FURUTA Department of Poultry Husbandry, University of California, Berkeley, California (Received for publication August 25, 1969) PREVIOUS work has shown that ingestion of raw soybean () diets caused greater decreases in proteases (Lepkovsky et al., 1965) and amylase (Lepkovsky et al., 1966) in the chicken pancreas than did ingestion of heated soybean () diets. Those decreases were accompanied by larger amounts of amylase in the intestinal contents. The amounts of proteases in the intestinal contents in the chickens fed diet were masked by the complexing of trypsin with the trypsin inhibitors in the. In the study reported here, lipase activity was measured in: 1) pancreas and intestinal and cecal contents of intact chickens; and 2) intestinal contents, and intestinal effluent collected through ileostomies, of chickens fed and diets. EXPERIMENTAL PROCEDURE Experiment 1. Intact chickens. Five-weekold Single Comb White Leghorn female chickens weighing 400 to 500 gm. were fed or diets*, with and without methionine supplement, for 6 weeks. At autopsy, the pancreases and the contents * Composition of diets in Lepkovsky et al. (1965). of the upper half of the small intestine, lower half of the small intestine and ceca were collected, frozen and lyophilized. Experiment 2. Chickens with ileostomies. Single Comb White Leghorn female chickens 5 weeks old were placed on or diets* either with or without methionine. After the chickens had been on the diets for 6 weeks, ileostomies were performed. When the chickens had recovered sufficiently and were eating well, they were fasted for 18 hours. The chickens on the diet were divided into 5 groups of 5 each; the control group was fasted for an additional 5 hours ( 23 hours); the remaining 4 groups were each offered one meal, for 5 hours, of diet, diet plus methionine, diet, or diet plus methionine. These are referred to as unadapted chickens. The chickens that were fed the experimental diets ( diet plus methionine, diet, or diet plus methionine) for 6 weeks were treated as follows: Food was withheld from half of each group for another 5 hours ( 23 hours); these acted as the fasting group. The remaining chickens were offered the diet for 5 hours. At the end of the 5-hour

2 LIPASE AND SOYBEANS 193 feeding period all the fasted and fed chickens were killed and examined. These are referred to as adapted chickens. The intestinal contents expelled through the ileostomy (intestinal effluent) during the 5-hour feeding period were combined with the contents remaining in the intestine, and were then frozen and lyophilized. The pancreases were removed, frozen and lyophilized. The experiment was repeated with the difference that only unadapted chickens were used, and the intestinal contents and intestinal effluent were collected separately and analyzed separately. The lipase activity was estimated in these samples by incubating them according to the method of Balls et al. (1937). The fatty acids released were determined by Dole's (1956) procedure. The incubation mixture consisted of an emulsion made up of a 1:1 mixture of olive oil and 16.5% gum arabic solution, 0.1 ml.; 5% sodium taurocholate, 0.5 ml.; 0.1 M CaCl 2, 0.5 ml; 0.6 M NH 4 C1-NH 3 buffer (ph 8.5), 0.3 ml.; H 2 0, 0.1 ml.; sample, 0.5 ml.; volume 2.0 ml. This volume was divided into two 1-ml. portions. Lipase action was stopped in one portion Unadapted chickens: M M Adapted chicken: M M (control) by the immediate addition of the extraction mixture; the other portion was incubated at 37 C. for 20 minutes before the extraction mixture (Dole, 1956) was added. When available, samples of pancreases and intestinal contents were analyzed for lipase activity without the use of bile salts. Addition of bile salts to some samples of intestinal contents increased, had no effect, or inhibited lipase activity. Accordingly, 56 random samples of pancreatic juice obtained from fistulated pancreatic ducts of 7 unanesthetized chickens were assayed for lipase with and without the addition of bile salts. RESULTS Intact chickens. Levels of lipase were lower in pancreases from unadapted chickens fed diets than in those of chickens fed diets, suggesting a greater secretion of lipase in the pancreatic juice of the chickens fed the diets (Table 1). This was not reflected in the contents of the upper halves of the intestines since those of chickens fed diets showed a little more lipolytic activity than did those of the chickens fed diets. As intestinal contents moved caudally, levels TABLE 1.Lipase activity of pancreases, and solids and lipase activity of upper and lower intestinal contents and cecal contents of unadapted chickens and of chickens adapted to heated () or raw () soybean diets with and without methionine (M) for 6 weeks solids Lipase activity* Intestinal contents after feeding Upper half Lipase solids Lower half Lipase Cecal contents after feeding solids Lipa.se * Expressed as Meq. free fatty acid released in 20 minutes at 37 C. sample.

3 194 S. LEPKOVSKY AND F. FURUTA of lipase decreased progressively, and were lowest in the cecal contents of the chickens fed diet. The reverse was true in the chickens fed diets; levels of lipase increased progressively and were highest in the cecal contents (Table 1). es of fasting chickens adapted to diets showed increased weight over those of chickens maintained on diets (Table 1), in agreement with previous work (Chernick et al., 1948). In contrast to chickens not adapted to diets, those so adapted showed higher pancreas lipase values following feeding than did corresponding chickens fed diets. Lipolytic activity in the upper intestinal contents of the chickens fed diets was somewhat higher than that in chickens fed diets (Table 1). As the intestinal contents moved caudally, the results obtained were similar to those found in the unadapted chickens (Table 1). Chickens with ileostomies. Responses of the pancreases of ileostomized chickens (Tables 2 and 3) to or diets were similar to those obtained with intact chickens (Table 1). Lipase activity in the intestinal contents or intestinal effluent was higher in chickens fed diets than in those fed diets (Tables 2 and 3), and no differences were seen between adapted and unadapted chickens. On a basis, little difference was seen between lipase activity in intestinal contents collected from the intestine and that in intestinal effluent (Table 3). lipase activity was similar in intestinal contents and intestinal effluent of chickens fed diet. In chickens fed the diets, lipase activity was higher in intestinal effluent than in the intestinal contents because of larger amounts of solids in the effluent. Bile salts in the determination of lipase activity. The results are shown in Table 4. Bile salts increased lipase values moderately (0 to 20%) in the pancreases of chickens fed either or, and to a greater extent in many samples of intestinal contents, but the latter values fluctuated widely. Lipase activity was inhibited in two cases (Table 4). In each of seven chickens, bile salts increased lipolytic activity of some sam- TABLE 2.Lipase activity in pancreases and in collected intestinal contents mixed with intestinal effluent of adapted and unadapted chickens fed heated () or raw () soybean diets with and without methionine (M) s Unadapted chickens: M M Adapted chickens: M M S Lipas ie activity* Intestinal contents mixed with intestinal effluent * Expressed as Meq. free fatty acid released in 20 minutes at 37 C. sample.

4 LIPASE AND SOYBEANS 195 TABLE 3.Lipase activity of the pancreases of fasted and fed chickens and of the lipase activity of collected intestinal contents and of intestinal effluent of unadapted chickens fed heated () or raw () soybean diets with and without methionine (M) Lipase activity* Intestinal contentsintestinal effluent M M pergm Intestinal T?fa «t Intestinal,,«, t Effluent Effluent contents contents ; Expressed as Meq. free fatty acid released in 20 minutes at 37 C. sample. pies of pancreatic juice, decreased it in some, and in still others, had no effect (Table 5). The samples were collected at different times from the same chicken. DISCUSSION The difficulty in interpreting the data obtained in studies on the nutritional value of and diets may be illustrated by the results obtained with chickens fed one meal of diet. Levels of lipase were lower in pancreases of these chickens than in those of chickens fed diet. This finding suggested that more lipolytic activity was secreted into the intestine after the diet. However, instead of more lipolytic activity in the contents of the upper halves of the small intestines of the chickens fed diet, somewhat less was found. It is important to keep in mind that lipase activity of the contents of the upper half of the intestine is most representative of the lipase activity that enters the small in- TABLE 4.Lipase activity, determined with () and without () bile salts, in pancreases and in contents of the intestine and ceca of individual chickens fed heated () and raw () soybean diets : : Upper Lipase activity* Intestine Lower Ceca * Expressed as Meq. free fatty acid released in 20 minutes at 37 C. sample.

5 196 S. LEPKOVSKY AND F. FURUTA TABLE 5.Lipase activity, determined with () and without () bile salts, in selected samples of pancreatic juice. All samples of pancreatic juice luted under each of the seven chickens were collected at different times from the same chicken Lip* ase activity* C C C C C C C * Expressed as Meq. free fatty acid released in 20 minutes at 37 C sample. testine with the pancreatic juice. The lipase activity of the intestinal contents is less representative because changes occur in the contents of the lower half (Table 1). An alternative explanation for the marked decreases in lipolytic activity in the pancreases of chickens fed the diet is that the synthesis of lipase in the pancreas is impaired by the diet during the feeding period. Final judgment cannot be made without measurements of synthesis of lipase during feeding. After chickens were adapted to the diet by chronic feeding, these differences disappeared. Such conditioning presumably caused changes in synthesis and release of lipase from the pancreas. The data indicate that lipase lacks stability in the intestinal contents of chickens fed diet, because there was less lipase activity in cecal contents than in the contents of the upper half of the intestine of those chickens. This conclusion is supported by the balance sheet (Table 6) which compares the calculated amounts of lipase "released" from the pancreas with those recovered from the intestinal contents during a 5-hour feeding period (Table 2). The amount of lipase "released" from the pancreas represents the difference between the amount of lipase at fasting and that found after feeding, and takes no account of synthesis during the feeding period. The intestinal contents of chickens fed diet contained more lipase "released" from the pancreas than did the intestinal contents of chickens fed diet. It is possible that the smaller "decreases" of lipase activity in the pancreases of chickens fed diets (Table 2) were due to higher rates of synthesis of lipase. In that case, the actual amount of lipase entering their intestines was greater than the calculated value, and exaggerated the apparent instability of lipase in intestinal contents of chickens fed diet. In contrast, lipase activity appeared stable in the intestinal and cecal contents of chickens fed diet, possibly a result of the diet's stabilizing action or because of excessive secretion of lipase into the intestinal lumen. Considerations discussed above appear to rule out excessive secre- TABLE 6.Balance sheet of lipase activity in pancreases and in corresponding intestinal contents of adapted chickens at fasting and after eating heated () or raw () soybean diets for 5 hours M M lipase activity- Decrease after eating for 5 hours Recovery in intestinal contents intestinal effluent after feeding for 5 hours

6 LIPASE AND SOYBEANS 197 tion of lipase from the pancreas, but the diet could have stimulated excessive secretion of lipase from the intestine. Pavlov and his students (1910) showed that intestinal juice (succus entericus) contains lipase. A small amount of intestinal juice was obtained from Thiry- Vella fistulas of the chicken, and presence of lipase was established. This preliminary observation could not be explored in the present study since the necessary longlasting Thiry-Vella fistulas could not be obtained because, unlike rats, chickens develop obstructions at the anastomoses of the intestine. An indirect method was used to determine whether appreciable amounts of non-pancreatic lipase were present in the intestinal contents of chickens. Pavlov (1910) found that lipase derived from intestinal juice of the dog did not require bile salts for its activation. A study of the action of bile salts upon lipolytic activity of chicken pancreatic juice showed that bile salts activated some samples and inhibited others. Different samples of pancreatic juice from the same chicken, collected at different times, were either activated or inhibited by bile salts or were left unchanged. If we assume that the chemical nature of lipase from different samples of pancreatic juice was the same, the environment of the intestinal lumen was responsible not only for the stability or lability of lipase but also for the differing action of bile salts upon its activity. In any case, pancreatic lipase could not be differentiated from intestinal lipase by the use of bile salts. SUMMARY Lipase activity was measured in: 1) pancreases of intact chickens and in pancreases of chickens with ileostomies; 2) intestinal contents from intestines of intact and operated chickens; 3) intestinal contents voided through an ileostomy; and 4) cecal contents from intact chickens. After one feeding of diet, the level of lipolytic activity in the pancreases of the chickens was reduced to about onehalf of that found at fasting, and was in marked contrast to the small decreases of lipolytic activity in the pancreases of chickens fed diet. After adaptation to the diet for 6 weeks, the converse occurred; more lipase activity remained in the pancreases of chickens fed diet than in those of chickens fed diet. The lipase activity of the contents of the upper half of the intestine was similar in chickens fed one meal of or diet; consequently, it did not reflect the differences in lipolytic activity noted above in the pancreases. As the intestinal contents moved caudally, lipase activity decreased in chickens fed diet, suggesting instability of lipase; in contrast, lipase activity increased in chickens fed diet. This increase could have been due to the stability of lipase or to excessive secretion of lipase into the intestinal lumen. Limited work established the presence of lipase in intestinal juice collected from a Thiry-Vella fistula in the intestines of chickens. Attempts to determine the action of or diets on the secretion of lipase in intestinal juice failed due to technical difficulties. Attempts to differentiate between pancreatic and intestinal lipase with the use of bile salts failed because bile salts increased lipolytic activity of some samples of pancreatic juice, was without effect on others, and inhibited lipolytic activity in still others. ACKNOWLEDGMENTS This investigation was supported in part by USP research grants A-1804

7 198 S. LEPKOVSKY AND F. FURUTA and NB-7585 from the National Institutes of Health, and GB-242 and GB from the National Science Foundation. REFERENCES Balls, A. K., M. B. Mattlock and J. W. Tucker, The hydrolysis of glycerides by crude pancreas lipase. J. Biol. Chem. 122: Chernick, S. S., S. Lepkovsky and I. L. Chaikoff, A dietary factor regulating the enzyme content of the pancreas: changes induced in size and proteolytic activity of the chick pancreas by the ingestion of raw soy-bean meal. Am. J. Physiol. 155: THE silicates, more commonly known as bentonite, kaolins, vermiculite and other clays, have been used in animal feeds and industrially for many years. They are used in feeds principally because of their ability to enhance pellet durability. As a group, they often are referred to as "pellet binders". Eventually, they may be widely used to control "watery droppings" in poultry. The highly adsorptive characteristic of binders led to experiments in this area (Quisenberry and Bradley, 1964). These workers found that sodium bentonite at both the 2.5 and 5.0% dietary levels significantly reduced the moisture content of the droppings of cage layers. Similarly, they have been proposed by some for the prevention of "loose droppings" in broilers where such 1 Mississippi Agricultural Experiment Station Journal Article No Silicates in Broiler s 1 Dole, V. P., A relation between nonesterified fatty acids in plasma and the metabolism of glucose. J. Clin. Invest. 35: Lepkovsky, S., F. Furuta, T. Koike, N. Hasegawa, M. K. Dimick, K. Krause and F. J. Barnes, The effect of raw soya beans upon the digestion of proteins and upon the function of the pancreas of intact chickens and of chickens with ileostomies. Brit. J. Nutr. 19: Lepkovsky, S., T. Koike, M. Sugiura, M. K. Dimick and F. Furuta, Pancreatic amylase in chickens fed on soya-bean diets. Brit. J. Nutr. 20: Pavlov, I. P., The Work of the Digestive Glands. Charles Griffin & Co., London. ELBERT J. DAY, R. D. BUSHONG, JR. AND BEN C. DILWORTH Department of Poultry Science, Mississippi State University, P. O. Box 5188, State College, Mississippi (Received for publication August 25, 1969) conditions cause increased condemnation in dressing plants. The highly adsorptive nature of the silicates led to studies to determine whether or not they adversely affect nutrient availability (Briggs and Spivey, 1954; Laughland and Phillips, 1954). These workers found that bentonite could create a vitamin A deficiency under certain conditions where purified diets were employed. However, recent data obtained with practical diets indicate that silicates enhanced feed efficiency or caloric efficiency (Cage layers; Quisenberry and Bradley, 1964: Turkeys; Almquist et ah, 1967: Broilers; Kurnick and Reed, 1960; Ousterhout, 1966). Several of the above research groups found that the silicates may act in slowing feed passage time through the digestive system. This slowing action has been postulated to be the explanation as to how they improve nutrient utilization by longer passage time

Animal Nutrition. Chapter 41. Biology Eighth Edition Neil Campbell and Jane Reece. PowerPoint Lecture Presentations for

Animal Nutrition. Chapter 41. Biology Eighth Edition Neil Campbell and Jane Reece. PowerPoint Lecture Presentations for Chapter 41 Animal Nutrition PowerPoint Lecture Presentations for Biology Eighth Edition Neil Campbell and Jane Reece Lectures by Chris Romero, updated by Erin Barley with contributions from Joan Sharp