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Long-term effect of feeding a new fiber fraction from soybean to zucker fatty rats
NUTRITION RESEARCH, Vol. 5, pp. 1161-1164, 1985 0271-5317/85 $3.00 + .00 Printed in the USA. Copyright (c) 1985 Pergamon Press Ltd. All rights reserved.
LONG-TERMEFFECTOFFEEDINGANEWFIBER FRACTION FROM SOYBEAN TO ZUCKERFATTY RATS Zecharia Madar, Ph.D. Department of Agricultural Biochemistry Faculty of Agriculture The Hebrew UniversitY of Jerusalem P.O. Box 12 Rehovot 76100, Israel
ABSTRACT
Feeding for a 14-week period, of a diet containing soybean dietary fiber to Zucker fatty (fa/fa) rats decreased plasma glucose and insulin levels, while plasma triglyceride and cholesterol levels remained unchanged. The triglyceride and cholesterol levels in liver and epididymal fat-pads were lower in fiber-fed rats compared to those fed without fiber. Fecal weight was higher and transit time was also shorter in these rats. Soybean dietary fiber decreased the protein fecal excretion. K_e_yWords: fiber, soybean, glucose, tolerance, fatty rats.
INTRODUCTION
It has been shown that lack of dietary fiber contributes to hyperinsulinaemia, obesity and Type II diabetes (I). Most fiber sources when incorporated into baked products in large qnantites have proven to be inedible altbough soybeans are a well-known source of vegetable protein ingested by a high-percentage of populations. Soybean dietary fiber is highly acceptable to human subjects (2) due mainly to its ease in usage without effecting palatability, Our previous study demonstrated that soybean, containing 35% fiber improved the glucose tolerance test and decreased plasma lipid levels in ob/ob mice (3). We undertook the present study to determine whether or not inclusion of new fiber preparation from soybean in diets of Zucker fatty (fa/fa) rats would affect plasma factors, such as glucose, insulin, lipids, and bowel function.
MATERIALS AND METHODS
Two groups of ten. young-male, Zucker fa/fa (fatty) rats were individually housed in stainless steel cages in a controlled enviromnent (22+_2~ Food and water were provided ad libitum. One group of rats was fed a fiber-free diet and the second group a fiber-enriched diet containing soybean as a source of fiber (75%) and protein (20%). Both groups were fed these diets for 14 weeks and were weighed weekly. Diet composition is shown in Table I. The diets were isocaloric (382 kcal) and the level of available carbohydrate was similar. The dietary fiber content in the soybean was determined according to Schweizer and Wursch (4). The rats were fasted for 12 hours prior to the oral glucose tolerance test at the start of the study, and after 8 and 14 weeks of consuming the diets. A glucose solution (0.3 g/100 g body weight) was administered orally by an intragastric tube and blood samples were collected from the tail veins. Plasma glucose was measured by the glucose oxidase method using a Beckman analyzer. Plasma immunoreactive insulin concentration was determined by the standard RIA using 1161
1162
Z. MADAR
125I-insulin (Novo, Copenhagen; Denmark) and guinea pig anti-porcine insulin (Serono Diagnostic Hypolab). Carmine red dye (1 ml; 0.1Z) was intubated for all the rats on days 30, 60 and I00 and the intestinal transit time was measured when the first colored fecal excretion appeared. TABLE I The Composition of Isocaloric Diets Ingredients*
Fiber-Free Diet g: KcaI
Corn Starch Glucose Casein ~ Soybean # Soybean Oil Total
61.2 5.0 18.0 5.0 93.4
* #
244.8 20.0 72.0 45.0 381.8
Fiber-Enriched Diet g Kcal 61.2 5.0 13.0 25.04 5.0 113.35
244.8 20.0 52.0 20.0 45.0 381.8
Mineral mixture 3.5 g, vitmnins 0.2 g, choline chloride 0.15 g and methionine 2 g. Contains 75% dietary fiber, 20%protein, 4% ash and 1% others.
Three-day fecal samples (collected every half hour) of each rat were pooled and dried to constant weight at I05~ This was determined three times during the experimental period after 30, 60 and 100 days on diet. The total amount (expressed as percentage) of water was calculated by the difference in the fecal samples' weight before and after the fecal samples were dried. Protein content in the fecal samples was determined according to the method of Kjeldahl. using an automatic analyzer. A t the conclusion of the study~ the rats were fasted for 12 hours and were anaesthetized with ether, the abdomen was opened and blood samples were collected from the vena cava for determination of triglycerides and total cholesterol. Aliquots of liver and epididymal fat pads were removed and extracted with chloroform:methanol (2:1) and assayed for triglyceride and cholesterol content. Plasma, liver and fat tissue triglycerides content were determined by the acteylacetone method (Sigma Chemical Co., St. Louis, MO), and cholesterol, measured colormetrically with the improved Liebermann-Buchard reagent (Sigma Chemical Co.) using serum as a standard* Data are presented as means +_SEM, Statistical analysis was performed by paired student's t t e s t . RESULTS
Oral glucose tolerance t e s t was performed a t week 8. No s i g n i f i c a n t difference i n glucose levels was shown between ~the two groups. The fasting and response insulin levels were not different (data not shown), A n additional glucose tolerance test was performed at week 14 of the experiment (FIG I). The change in glucose levels were notably lower following glucose loading a t 60, 120 and 180 minutes i n the r a t s fed the fiber-enriched diet than those fed the f i b e r - f r e e diet. The r i s e in plasma i n s u l i n a t 30 and 60 minutes (FIG 1B), was generally smaller in the f i b e r - e n r i c h e d group and reached a s i g n i f i c a n t difference a t 120 and 180 minutes (p<0.05) following glucose loading. The soybean dietary f i b e r sUpplement had only a s l i g h t influence, but not s i g n i f i c a n t on the plasma cholesterol and t r i g l y c e r i d e s I e v e i s (Table 2). Cholestero! and triglyceride levels in the liver tissue were significantly lower in the rats fed the fiber-enriched diets. Cholesterol levels in epididymal fat pads were also lower in the rats fed the fiber-enriched diet. However, the triglyceride levels were identical in both groups 9 Rats fed a f i b e r - f r e e d i e t excreted 0.81_+0.21 g/day of feces (expressed by weight) vs. 1.43+0.33 i n r a t s fed the fiber-enriched diet. The fecal water (Z) was 16.60+--2.91 vs. 26§ and fecal protein excretion (~) was 29~56+_I,16vs. 16.33_+0,64. The ,,~,n tr,n~t time (hr) was 13_+0.50 in rats fed without fiber and 9_+0.75 with fiber.
SOYBEAN FIBER IN ZUCKERRATS
1163
FIG i Changes inglucoseand insulin levels (from fasting levels) in fa/fa obese ratsfed fiber-free (~--e) or fiber-enriched diet (o--o) for 14 weeks. The rats weredosed with glucose solution and the plasma glucose and insulin were determined at 0, 30, 60, 120and 180minutes. The values are expressed as means+SEM (n=10) after subtraction of zerovalue from each time. A. Change in plasma glucose levels. B. Change in insulin levels.
I
I
I
i
,~8 P"O 05
|
T
I:~0.05
o
60
I
120 TIME ~nin)
I
~0
TABLE 2
Effect of Soybean Dietary Fiber on Plasma, L i v e r ~ n d Epididymal Fat Pads Cholesterol and Triglycerides Fiber-Free Diet Plasma (mg/dl) Total Cholesterol Triglycerides Liver (mg/100g) Cholesterol Triglycerides Epididymal Fat Pads (m~/100g) Cholesterol Triglycerides
741• 378• 452+55* 1713~290" 273__+49.0* 312--+0.97
Fiber-Enriched Diet
701• 354• 173• 892+_86.0 108+_15.4 314--+0.65
Values represent mean +SEM, n=10 # Determined at conclusion of the study t14 weeks on dzets) and after 12 h of fasting. * Significantly different from fiber-enriched diet at P<0~5. DISCUSSION
Soybean dietary fiber fraction (75%) ingestion in genetically Zucker (fa/fa) rats had
1164
Z. MADAR
no effect on the glucose levels after 8 weeks of feeding. However, maintaining the rats on a fiber-enriched diet for 14 weeks improved the oral glucose tolerance test at 60, 120 and 180 minutes following glucose loading. We cannot fully explain at this stage why the plasma glucose levels significantly decreased only after 8 weeks. However, these results indicated that adaptiou to fiber ingestion is important for its effectiveness in improving glucose metabolism. Thus, adaptation involves changes in gastrointestinal tract structure which leads to impairment in glucose passing across the intestine and in secretion of GIP (7). Together, these lead to decreased glucose and insulin levels. However, it is expecte d that gut adaption to liver should occur after 8 weeks, although no changes in glucose levels were observed. The soybean fiber contains pectins, gulactomannas and arabinogalactan with high viscosity. These polysaccharides slow gastric emptying and glucose absorption (2). Zucker fatty rats fed fiber had lower levels of cholesterol and triglycerides in the liver while there was no difference in plasma cholesterol levels compared to non-fiber-fed rats. This could be the result of cholesterol and bile acid being bound by fiber in the intestine and/or by increased fecal steroid excretion. Soybean dietary fiber also decreased the cholesterol level in epididymal fat pads which may not be attributed to a decrease in plasma cholesterol. It was previously reported that cholesterol levels in serum do not reflect rates of biosynthesis or degredation of cholesterol (6). It is also possible that changes in plasma cholesterol occur only when the initial cholesterol levels are high (6). Soybean dietary fiber induced the expected increase in fecal output, and transit time was shortened. These effects, together with a higher water-holding capacity in feces of rats fed fiber, may removed glucose from the intestinal tract and contribute to lowering the glucose level, in the present study, the loss of protein excretion in rats fed soybean dietary fiber was significantly less than in rats fed without fiber. These results may suggest that soybean dietary fiber improved protein utilization in genetically obese rats which generally have impaired protein metabolism (7). Our prior study with ob/ob mice (3) and in other studies with normal animals (7) show less protein retention with fiber ingestion. Thus, the effect of 75% soybean fiber fraction on protein retention needs further clarification. However, the fiber excretion was not determined, therefore, the possibility of the dilution of feces by fiber can not be excluded. In conclusion, this study provides some evidence for the effectiveness of new fraction of soybean dietary fiber on carbohydrate and lipid metabolism in obese rats.
AC~OWLEDGEm~_ The author wishes to thank Ms. Nava Cohen for her excellent technical assistance, and Ms. Nadine Daleo for her secretarial assistance. This work was supported by grants from '~oor Mason-Hayes," Ashdod, israel. REFERENCES
I. Staub P,AII, Laveille CA. Dietary Fiber and Obesity. In: Vahouny GV and Kritchevsky D eds. Dietary Fiber in Health and Disease. Plenum Press, New York, 1982:139-49. 2. Schweizer TF. Bekhechi AR, Koellreutter B, Reimann S, Ponnetta D, Brown Bit Metabolic Effects of Dietary Fiber From Dehulled Soybean in Humans. Am. J. Clin. Nutr. 1983;
38:1-11. 3. Medar Z, Reyzak O, Trostler N. Reduction of Glucose and Insulin Levels in ob/ob obese Mice Fed Soybean Dietary Fiber. Nutr. Res. 1985; 5:305-315. 4. Schweizer THE. Wursch P. Analysis of Dietary Fiber. J. Sci. Food Agr. 1978; 28:148-54. 5. Leeds AIL Modification of Intestinal Absorption by Dietary Fiber and Fiber Components. In: Vohouny GV and Kritchevsky D eds. Dietary-Fiber in Health and Disease, Plenum Press, New York, 1982; 53-71. 6. Terphsa AHM, West CE. Fennis JCM, Schouten JA, Veto AE. Hypucholesterolaemic Effect of Dietary Soy Protein Versus Casein in Rhesus Monkeys (Macaca mulatta). Am. J. Clin. Nutr. 1983; 39:1-7. 7. Deb S, Martin RJ, Hershberger EV. Maintenance Requirement and Energetic Efficiency of Lean and Obese Zucker Rats. J. Nutr. 1976; 106:191-97.
Accepted f o r p u b l i c a t i o n August 25, 1985.
LONG-TERMEFFECTOFFEEDINGANEWFIBER FRACTION FROM SOYBEAN TO ZUCKERFATTY RATS Zecharia Madar, Ph.D. Department of Agricultural Biochemistry Faculty of Agriculture The Hebrew UniversitY of Jerusalem P.O. Box 12 Rehovot 76100, Israel
ABSTRACT
Feeding for a 14-week period, of a diet containing soybean dietary fiber to Zucker fatty (fa/fa) rats decreased plasma glucose and insulin levels, while plasma triglyceride and cholesterol levels remained unchanged. The triglyceride and cholesterol levels in liver and epididymal fat-pads were lower in fiber-fed rats compared to those fed without fiber. Fecal weight was higher and transit time was also shorter in these rats. Soybean dietary fiber decreased the protein fecal excretion. K_e_yWords: fiber, soybean, glucose, tolerance, fatty rats.
INTRODUCTION
It has been shown that lack of dietary fiber contributes to hyperinsulinaemia, obesity and Type II diabetes (I). Most fiber sources when incorporated into baked products in large qnantites have proven to be inedible altbough soybeans are a well-known source of vegetable protein ingested by a high-percentage of populations. Soybean dietary fiber is highly acceptable to human subjects (2) due mainly to its ease in usage without effecting palatability, Our previous study demonstrated that soybean, containing 35% fiber improved the glucose tolerance test and decreased plasma lipid levels in ob/ob mice (3). We undertook the present study to determine whether or not inclusion of new fiber preparation from soybean in diets of Zucker fatty (fa/fa) rats would affect plasma factors, such as glucose, insulin, lipids, and bowel function.
MATERIALS AND METHODS
Two groups of ten. young-male, Zucker fa/fa (fatty) rats were individually housed in stainless steel cages in a controlled enviromnent (22+_2~ Food and water were provided ad libitum. One group of rats was fed a fiber-free diet and the second group a fiber-enriched diet containing soybean as a source of fiber (75%) and protein (20%). Both groups were fed these diets for 14 weeks and were weighed weekly. Diet composition is shown in Table I. The diets were isocaloric (382 kcal) and the level of available carbohydrate was similar. The dietary fiber content in the soybean was determined according to Schweizer and Wursch (4). The rats were fasted for 12 hours prior to the oral glucose tolerance test at the start of the study, and after 8 and 14 weeks of consuming the diets. A glucose solution (0.3 g/100 g body weight) was administered orally by an intragastric tube and blood samples were collected from the tail veins. Plasma glucose was measured by the glucose oxidase method using a Beckman analyzer. Plasma immunoreactive insulin concentration was determined by the standard RIA using 1161
1162
Z. MADAR
125I-insulin (Novo, Copenhagen; Denmark) and guinea pig anti-porcine insulin (Serono Diagnostic Hypolab). Carmine red dye (1 ml; 0.1Z) was intubated for all the rats on days 30, 60 and I00 and the intestinal transit time was measured when the first colored fecal excretion appeared. TABLE I The Composition of Isocaloric Diets Ingredients*
Fiber-Free Diet g: KcaI
Corn Starch Glucose Casein ~ Soybean # Soybean Oil Total
61.2 5.0 18.0 5.0 93.4
* #
244.8 20.0 72.0 45.0 381.8
Fiber-Enriched Diet g Kcal 61.2 5.0 13.0 25.04 5.0 113.35
244.8 20.0 52.0 20.0 45.0 381.8
Mineral mixture 3.5 g, vitmnins 0.2 g, choline chloride 0.15 g and methionine 2 g. Contains 75% dietary fiber, 20%protein, 4% ash and 1% others.
Three-day fecal samples (collected every half hour) of each rat were pooled and dried to constant weight at I05~ This was determined three times during the experimental period after 30, 60 and 100 days on diet. The total amount (expressed as percentage) of water was calculated by the difference in the fecal samples' weight before and after the fecal samples were dried. Protein content in the fecal samples was determined according to the method of Kjeldahl. using an automatic analyzer. A t the conclusion of the study~ the rats were fasted for 12 hours and were anaesthetized with ether, the abdomen was opened and blood samples were collected from the vena cava for determination of triglycerides and total cholesterol. Aliquots of liver and epididymal fat pads were removed and extracted with chloroform:methanol (2:1) and assayed for triglyceride and cholesterol content. Plasma, liver and fat tissue triglycerides content were determined by the acteylacetone method (Sigma Chemical Co., St. Louis, MO), and cholesterol, measured colormetrically with the improved Liebermann-Buchard reagent (Sigma Chemical Co.) using serum as a standard* Data are presented as means +_SEM, Statistical analysis was performed by paired student's t t e s t . RESULTS
Oral glucose tolerance t e s t was performed a t week 8. No s i g n i f i c a n t difference i n glucose levels was shown between ~the two groups. The fasting and response insulin levels were not different (data not shown), A n additional glucose tolerance test was performed at week 14 of the experiment (FIG I). The change in glucose levels were notably lower following glucose loading a t 60, 120 and 180 minutes i n the r a t s fed the fiber-enriched diet than those fed the f i b e r - f r e e diet. The r i s e in plasma i n s u l i n a t 30 and 60 minutes (FIG 1B), was generally smaller in the f i b e r - e n r i c h e d group and reached a s i g n i f i c a n t difference a t 120 and 180 minutes (p<0.05) following glucose loading. The soybean dietary f i b e r sUpplement had only a s l i g h t influence, but not s i g n i f i c a n t on the plasma cholesterol and t r i g l y c e r i d e s I e v e i s (Table 2). Cholestero! and triglyceride levels in the liver tissue were significantly lower in the rats fed the fiber-enriched diets. Cholesterol levels in epididymal fat pads were also lower in the rats fed the fiber-enriched diet. However, the triglyceride levels were identical in both groups 9 Rats fed a f i b e r - f r e e d i e t excreted 0.81_+0.21 g/day of feces (expressed by weight) vs. 1.43+0.33 i n r a t s fed the fiber-enriched diet. The fecal water (Z) was 16.60+--2.91 vs. 26§ and fecal protein excretion (~) was 29~56+_I,16vs. 16.33_+0,64. The ,,~,n tr,n~t time (hr) was 13_+0.50 in rats fed without fiber and 9_+0.75 with fiber.
SOYBEAN FIBER IN ZUCKERRATS
1163
FIG i Changes inglucoseand insulin levels (from fasting levels) in fa/fa obese ratsfed fiber-free (~--e) or fiber-enriched diet (o--o) for 14 weeks. The rats weredosed with glucose solution and the plasma glucose and insulin were determined at 0, 30, 60, 120and 180minutes. The values are expressed as means+SEM (n=10) after subtraction of zerovalue from each time. A. Change in plasma glucose levels. B. Change in insulin levels.
I
I
I
i
,~8 P"O 05
|
T
I:~0.05
o
60
I
120 TIME ~nin)
I
~0
TABLE 2
Effect of Soybean Dietary Fiber on Plasma, L i v e r ~ n d Epididymal Fat Pads Cholesterol and Triglycerides Fiber-Free Diet Plasma (mg/dl) Total Cholesterol Triglycerides Liver (mg/100g) Cholesterol Triglycerides Epididymal Fat Pads (m~/100g) Cholesterol Triglycerides
741• 378• 452+55* 1713~290" 273__+49.0* 312--+0.97
Fiber-Enriched Diet
701• 354• 173• 892+_86.0 108+_15.4 314--+0.65
Values represent mean +SEM, n=10 # Determined at conclusion of the study t14 weeks on dzets) and after 12 h of fasting. * Significantly different from fiber-enriched diet at P<0~5. DISCUSSION
Soybean dietary fiber fraction (75%) ingestion in genetically Zucker (fa/fa) rats had
1164
Z. MADAR
no effect on the glucose levels after 8 weeks of feeding. However, maintaining the rats on a fiber-enriched diet for 14 weeks improved the oral glucose tolerance test at 60, 120 and 180 minutes following glucose loading. We cannot fully explain at this stage why the plasma glucose levels significantly decreased only after 8 weeks. However, these results indicated that adaptiou to fiber ingestion is important for its effectiveness in improving glucose metabolism. Thus, adaptation involves changes in gastrointestinal tract structure which leads to impairment in glucose passing across the intestine and in secretion of GIP (7). Together, these lead to decreased glucose and insulin levels. However, it is expecte d that gut adaption to liver should occur after 8 weeks, although no changes in glucose levels were observed. The soybean fiber contains pectins, gulactomannas and arabinogalactan with high viscosity. These polysaccharides slow gastric emptying and glucose absorption (2). Zucker fatty rats fed fiber had lower levels of cholesterol and triglycerides in the liver while there was no difference in plasma cholesterol levels compared to non-fiber-fed rats. This could be the result of cholesterol and bile acid being bound by fiber in the intestine and/or by increased fecal steroid excretion. Soybean dietary fiber also decreased the cholesterol level in epididymal fat pads which may not be attributed to a decrease in plasma cholesterol. It was previously reported that cholesterol levels in serum do not reflect rates of biosynthesis or degredation of cholesterol (6). It is also possible that changes in plasma cholesterol occur only when the initial cholesterol levels are high (6). Soybean dietary fiber induced the expected increase in fecal output, and transit time was shortened. These effects, together with a higher water-holding capacity in feces of rats fed fiber, may removed glucose from the intestinal tract and contribute to lowering the glucose level, in the present study, the loss of protein excretion in rats fed soybean dietary fiber was significantly less than in rats fed without fiber. These results may suggest that soybean dietary fiber improved protein utilization in genetically obese rats which generally have impaired protein metabolism (7). Our prior study with ob/ob mice (3) and in other studies with normal animals (7) show less protein retention with fiber ingestion. Thus, the effect of 75% soybean fiber fraction on protein retention needs further clarification. However, the fiber excretion was not determined, therefore, the possibility of the dilution of feces by fiber can not be excluded. In conclusion, this study provides some evidence for the effectiveness of new fraction of soybean dietary fiber on carbohydrate and lipid metabolism in obese rats.
AC~OWLEDGEm~_ The author wishes to thank Ms. Nava Cohen for her excellent technical assistance, and Ms. Nadine Daleo for her secretarial assistance. This work was supported by grants from '~oor Mason-Hayes," Ashdod, israel. REFERENCES
I. Staub P,AII, Laveille CA. Dietary Fiber and Obesity. In: Vahouny GV and Kritchevsky D eds. Dietary Fiber in Health and Disease. Plenum Press, New York, 1982:139-49. 2. Schweizer TF. Bekhechi AR, Koellreutter B, Reimann S, Ponnetta D, Brown Bit Metabolic Effects of Dietary Fiber From Dehulled Soybean in Humans. Am. J. Clin. Nutr. 1983;
38:1-11. 3. Medar Z, Reyzak O, Trostler N. Reduction of Glucose and Insulin Levels in ob/ob obese Mice Fed Soybean Dietary Fiber. Nutr. Res. 1985; 5:305-315. 4. Schweizer THE. Wursch P. Analysis of Dietary Fiber. J. Sci. Food Agr. 1978; 28:148-54. 5. Leeds AIL Modification of Intestinal Absorption by Dietary Fiber and Fiber Components. In: Vohouny GV and Kritchevsky D eds. Dietary-Fiber in Health and Disease, Plenum Press, New York, 1982; 53-71. 6. Terphsa AHM, West CE. Fennis JCM, Schouten JA, Veto AE. Hypucholesterolaemic Effect of Dietary Soy Protein Versus Casein in Rhesus Monkeys (Macaca mulatta). Am. J. Clin. Nutr. 1983; 39:1-7. 7. Deb S, Martin RJ, Hershberger EV. Maintenance Requirement and Energetic Efficiency of Lean and Obese Zucker Rats. J. Nutr. 1976; 106:191-97.
Accepted f o r p u b l i c a t i o n August 25, 1985.