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The Journal of Nutrition Vol. 128 No. 7 July 1998,
pp. 1156-1164
Azusawa Research Laboratories, Institute for Consumer Healthcare, Yamanouchi Pharmaceutical Company, Itabashi-ku, Tokyo 174, Japan and * Laboratory of Nutritional Biochemistry, Otsuma Women's University, Chiyodaku, Tokyo 102, Japan
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ABSTRACT |
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Abstract
Introduction
Methods
Results
Discussion
References
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The objective of this study was to examine the physiologic importance of undigested protein on cecal fermentation in rats fed a low (LAS) and high (HAS) amylose cornstarch. In Experiment 1, rats were fed diets containing LAS (655 g/kg diet) with one of four protein sources: casein, rice (RP), potato (PP) or soybean protein (SP) at 250 g/kg diet for 15 d. Apparent digestibilities of casein, RP, SP and PP were 96, 94, 93 and 92%, respectively. In rats fed the LAS diet with casein, acetate, propionate and succinate were the major cecal organic acids. The succinate pools in rats fed RP or SP were significantly lower than in those fed casein, whereas butyrate did not differ. Butyrate was significantly higher in rats fed PP, but succinate was the same as in rats fed casein. In Experiment 2, rats were fed diets containing HAS (200 g/kg diet) with one of the four protein sources at 250 g/kg diet for 10 d. HAS was substituted for the same amount of LAS. In rats fed the HAS diet, succinate was the major acid in rats fed casein; in rats fed RP or PP, however, the pools of this acid were significantly lower than in those fed casein, whereas butyrate was significantly higher in rats fed RP or PP. Fecal starch excretion was significantly lower in rats fed RP or PP than in those fed casein. In Experiment 3, rats were fed the casein-HAS diet with graded levels of PP (0, 10, 30, 50, 100 and 250 g/kg diet) for 14 d. The PP was substituted for the same amount of casein. Cecal butyrate was low in rats fed up to 100 g of PP/kg diet and then rose with 250 g of PP/kg diet. In Experiment 4, ileorectostomized rats were used and fed the same diets described in Experiment 3 for 9 d. The ileal starch/nitrogen ratio declined with increasing dietary PP, due solely to greater nitrogen excretion, whereas starch excretion was unaffected. In Experiment 5, rats were fed the casein-HAS diet with or without 60 g of artificial resistant protein/kg diet for 10 d. The resistant protein (apparent digestibility, 63%) was substituted for the same amount of casein. Rats fed the casein-HAS diet with resistant protein had significantly greater cecal butyrate and lower succinate than those fed the casein-HAS diet. These data show that large bowel fermentation of starch is altered by dietary protein. They support the hypothesis that nondigested protein, namely, resistant protein, may control fermentation efficiency as well as the fermentation profile of HAS, possibly as a result of a change in microflora through the change in the ratio of starch to nitrogen in the cecum.
KEY WORDS: resistant protein · high amylose cornstarch · cecal succinate · cecal butyrate · rats
Bacterial fermentation of dietary complex carbohydrates by the large bowel microflora has attracted considerable interest because the resulting short-chain fatty acids (SCFA)4 have potentially beneficial effects on large bowel physiology (Royall et al. 1990 Attention has shifted from non-starch polysaccharides (major components of dietary fiber) to starch as a major substrate for colonic fermentation. Starch (and the products of its digestion) that enters the large bowel is believed to be the most important fuel for human colonic microflora (Cummings and Macfarlane 1991). This resistant starch (RS) may have an additional advantage in that its fermentation appears to favor butyrate formation. Weaver et al. (1992) In our preliminary study, we noticed that the fermentation profile of HAS in rat cecum was drastically affected by dietary protein source. When casein was added as the sole protein source to the diet containing 200 g HAS/kg diet, the most predominant organic acid produced in the cecum was not SCFA, but succinate. On the other hand, when potato protein with lower digestibility was used in place of casein, the concentration of cecal succinate was decreased with a concomitant increase of SCFA, butyrate in particular. In turn, indigestible protein, namely, resistant protein, that enters the cecum may be involved in controlling the cecal fermentation of HAS in rats. The aim of this paper is to examine this hypothesis and to propose and discuss the physiologic importance of resistant protein on cecal fermentation in rats fed a HAS diet.
Materials.
Alkaline-extracted rice protein (RP, 122.6 mg nitrogen/g) was prepared as described previously (Morita et al. 1996
INTRODUCTION
Abstract
Introduction
Methods
Results
Discussion
References
). Of the three major SCFA, acetate, propionate and butyrate, the last-mentioned has attracted much attention. Colonocytes are believed to use SCFA as metabolic fuels with butyrate oxidation providing >70% of the oxygen consumed by colonic tissue (Roediger 1980a). Impaired utilization of SCFA may contribute to the pathogenesis of ulcerative colitis, which suggests that it may result from a deficiency of energy supply (Roediger 1980b). This view has been supported by Scheppach et al. (1992), who reported that direct infusion of butyrate into the colon leads to remission of distal ulcerative colitis.
have shown that in vitro, starch fermentation produces relatively more butyrate than the fermentation of non-starch polysaccharides. The data of Scheppach et al. (1988) and Englyst et al. (1987) from studies in humans support this view. However, Mallett et al. (1988) examined RS from two sources, high amylose cornstarch (HAS) and potato starch, in rats. They found that both increased cecal size and decreased cecal pH were most marked in rats fed potato starch. They found also that HAS increased large bowel SCFA but potato starch did not, suggesting that the substantial fall in pH elicited by potato starch might have been due to another organic acid such as lactate. Annison and Topping (1994) noted in pigs that RS in certain processed foods was an important contributor to large bowel SCFA but that both the molar ratios of the three major acids and their distribution along the colon differed with starch source (brown rice vs. navy beans). This would suggest that RS fermentation per se did not favor butyrate production; on the basis of supporting in vitro evidence (McBurney et al. 1988), they suggested that nitrogen availability might be an important factor in determining SCFA production. This may be of particular importance in the case of HAS fermentation, which seems to be relatively rapid (Topping et al., 1997). There are major two sources of nitrogen for the large bowel microflora, namely, urea (via the urease reaction) and protein escaping from the small intestine. Of these, the latter can be altered most readily by diet and thus would seem to be a possible means for modifying fermentation products.
MATERIALS AND METHODS
Abstract
Introduction
Methods
Results
Discussion
References
). Potato protein (PP, 122.6 mg nitrogen/g) was supplied by UnicoopJapan (Tokyo, Japan) and was prepared from potato juice by steam coagulation at pH 5.0-5.5 (Hisatsune, T., UnicoopJapan, personal communication). The chemical and amino acid compositions of these proteins have been reported previously (Morita et al. 1997). Casein (125.5 mg nitrogen/g) and soybean protein (SP, 126.2 mg nitrogen/g) were purchased from New Zealand Dairy Board (Wellington, New Zealand) and Fuji Oil (Osaka, Japan), respectively. The total dietary fiber concentrations of RP, PP and SP were determined by the method of Prosky et al. (1988) and were 11, 46 and 18 g/kg, respectively. The apparent digestibilities of casein, RP, PP and SP were determined by the method of Njaa (1959) under isonitrogenous condition in rats by using the following equation:
and were found to be 96, 94, 92 and 93%, respectively. Low amylose cornstarch (LAS, cornstarch W) was purchased from Nihon Shokuhin Kako (Tokyo, Japan) and HAS (Hi-maize) was purchased from Starch Australasia (Lane Cove, New South Wales, Australia). Because our previous studies (Ikai et al. 1997
) showed that in vivo, the RS value of HAS measured in ileorectostomized rats correlated with in vitro total dietary fiber value by the method of Prosky et al. (1988), the RS content of starches used in this study was tentatively defined as the amount of total dietary fiber. No RS was found in the LAS, whereas the HAS contained 250 g RS/kg. Artificial resistant protein was prepared in our laboratory by autoclaving freeze-dried egg white for 30 min at 100°C by the method of Yoshida et al. (1968). Its apparent digestibility was 63%.
Care of animals. Male Sprague-Dawley rats (Shizuoka Laboratory Animal Center, Hamamatsu, Japan) were housed individually in wire screen-bottomed stainless steel cages in a room with controlled temperature (23 ± 1°C) and lighting (lights on from 0800 to 2000 h). After adaptation to a casein-LAS diet (Table 1) for at least 5 d, rats were divided into groups on the basis of body weight and allowed free access to experimental diets and water. Body weight and food intake were recorded each morning before the diet was replenished.
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Dietary studies
Effects of casein, RP, PP and SP on cecal fermentation in rats fed a LAS diet (Experiment 1).
Twenty-four rats weighing 201-226 g were divided into four groups after acclimation and were allowed free access to diets containing casein, RP, PP or SP at 250 g/kg diet for 15 d. In this experiment, the sole source of carbohydrate was LAS, and the composition of each diet was the same as that of the casein-LAS diet (Table 1) except for protein sources. The RP, PP or SP was added to each diet at the expense of an equal amount of casein. Feces were collected for the last 3 d of the experimental period, freeze-dried and stored at 
20°C. At the end of the experimental period, rats were anesthetized with diethyl ether at 1300-1600 h, and the cecum was removed and weighed. The cecal contents were transferred to a 50-mL screw-capped glass tube and stored at 20°C until analysis. The cecal wall was flushed clean with ice-cold 0.15 mol/L NaCl, blotted dry on filter paper and weighed.
Effects of casein, RP, PP and SP on cecal fermentation in rats fed a HAS diet (Experiment 2). Twenty-four rats weighing 170-183 g were devided into four groups after acclimation and were allowed free access to diets containing casein, RP, PP or SP at 250 g/kg diet for 10 d. The basic composition of each diet was the same as that of the casein-LAS diet (Table 1) except for the protein and carbohydrate sources. High amylose cornstarch (200 g/kg diet) was added to each diet at the expense of an equal amount of LAS, i.e., the total amount of dietary starch was the same (655 g/kg diet) in all diets. The RP, PP or SP was added to each diet at the expense of an equal amount of casein. Fecal collection and analysis and sampling of cecal contents and their analyses were as described for Experiment 1.
Effects of the graded levels of PP on cecal fermentation in rats fed a HAS diet (Experiment 3). Thirty-six rats weighing 170-183 g were divided into six groups after acclimation, and were allowed free access to one of six diets containing 0, 10, 30, 50, 100 or 250 g of PP/kg diet. The basic composition of each diet was the same as that of the casein-LAS diet (Table 1) except for the protein and carbohydrate sources. High amylose cornstarch (200 g/kg of diet) was added to each diet at the expense of an equal amount of LAS, i.e., the total amount of dietary starch was the same (655 g/kg diet) in all diets. The graded levels of PP were added to each diet at the expense of an equal amount of casein so that the total amount of dietary protein was the same (250 g/kg diet), i.e., the 25% PP diet was free of casein. Sampling and analytical procedures were as described for Experiment 1.
Effects of the graded levels of PP on fecal excretions of nitrogen and starch in ileorectostomized rats fed a HAS diet (Experiment 4).
After acclimation to the casein-LAS diet, thirty-six rats weighing 200-250 g were subjected to an ileorectostomy in which the ilium is connected directly to the rectum as described previously (Nishimura et al. 1993). Rats subjected to this operation were not allowed food and water for the first 24 h after surgery; rats received daily intramuscular injections of 10 µL of Mycillin Sol [containing procaine penicillin G (200 g/L) and dihydrostreptomycin sulfate (250 g/L); Toyo Jozo, Shizuoka, Japan] on d 0-3 after surgery. They were then fed the casein-LAS diet (Table 1) for 7-10 d. Constant growth rates (5-7 g body weight gain/d) were achieved after 5 d. After postoperative recovery, the rats, weighing 250-300 g, were divided into six groups on the basis of body weight.
20°C.
Effects of an artificial resistant protein on cecal fermentation in rats fed a HAS diet (Experiment 5). Eighteen rats weighing 195-215 g were divided into three groups after acclimation and were allowed free access to one of three diets for 10 d. The composition of each diet was the same as that of the casein-LAS diet (Table 1) except for the protein and carbohydrate sources. Two groups were fed either LAS or HAS with 250 g of casein/kg diet. The third group was fed a HAS diet with 190 g of casein and 60 g of artificial resistant protein/kg diet. The latter addition was at the expense of casein so that the diets were balanced in protein concentration (250 g/kg diet). High amylose cornstarch (200 g/kg diet) was added to the diet at the expense of an equal amount of LAS, i.e., the total amount of dietary starch was the same in all diets (655 g/kg diet). Sampling and analytical procedures were as described for Experiment 1.
Analytical procedures.
After homogenization of cecal contents, a portion of homogenate was diluted with the same weight of distilled water, and cecal pH was measured with a compact pH meter (Model C-1, Horiba, Tokyo, Japan). Cecal organic acids (formate, acetate, propionate, isobutyrate, n-butyrate, isovalerate, n-valerate, citrate, malate, succinate and lactate) were measured by the internal standard method (Hoshi et al. 1994) using a HPLC (LC-6A, Shimadzu, Kyoto, Japan) equipped with Shim-pack SCR-102H column (8 mm i.d. × 30 cm long, Shimadzu) and an electroconductibity detector (CDD-6A, Shimadzu). Briefly, ~300 mg cecal contents were homogenized in 2 mL of 10 mmol/L sodium hydroxide solution containing 0.5 g/L crotonic acid as an internal standard and then centrifuged at 10,000 × g for 15 min. The supernatant obtained was applied to HPLC analysis. Fecal nitrogen was determined by the Kjeldahl method (Miller and Houghton 1945). Fecal starch was determined by using a Megazyme Total Starch Assay Kit (Megazyme Australia, Sydney, Australia) with a modification that involved preheating the samples in dimethylsulfoxide at 100°C for 30 min (Muir et al. 1995). The analysis of fecal neutral sterols was carried out according to the previous report (Morita et al. 1997). Calculation was according to the internal standard method using 5 
-cholestane. In these experiments, we defined neutral sterols tentatively as the sum of cholesterol and coprostanol.
Statistical analyses.
Data were analyzed by ANOVA; significant differences among means were separated by Duncan's multiple range test (Shibata 1974). When variances were not homogeneous by Bartlett test (Zar 1984), data were logarithmically transformed and transformed data were then analyzed by ANOVA followed by multiple comparison. Because variances for the coprostanol/cholesterol ratio varied among dietary groups even after logarithmic transformation of data, these results were presented as medians with range and then analyzed by Kruskal-Wallis ANOVA followed by Kolmogorov-Smirnov two-sample test (Zar 1984). Differences were considered significant at P < 0.05. The correlations among cecal pH, cecal tissue weight, cecal contents, cecal organic acids, and fecal starch and nitrogen, and dietary protein levels were analyzed by linear regression (Snedecor and Cochran 1967).
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RESULTS |
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Abstract
Introduction
Methods
Results
Discussion
References
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Effects of casein, RP, PP and SP on cecal fermentation in rats fed a LAS diet (Experiment 1). In this experiment, all rats were fed LAS with one of four sources of protein. There were no significant differences in food intake among the groups but body weight gain was significantly lower in rats fed PP than in those fed casein or SP (Table 2). The weights of cecal contents were significantly greater in rats fed PP than in the other three groups although the weight of cecal tissue did not differ among the groups (Table 2). Mean cecal pH values were 7 or greater in all groups (Table 2). pH was lowest in rats fed PP, highest in rats fed casein, and intermediate in the other two groups. Acetate was the major SCFA in cecal contents in all groups, and its pool size tended to be lower in rats fed casein than in the other three groups (P = 0.097, Table 2). That of propionate was unaffected by diet. The pool size of succinate equaled that of propionate in rats fed casein, whereas butyrate was very low in this group. Cecal succinate was significantly lower in rats fed SP or RP than in rats fed casein, but the pool sizes of butyrate did not differ in these three groups. The largest pool size of butyrate was found in rats fed PP, but succinate did not differ in this group from that in rats fed casein.
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Effects of casein, RP, PP and SP on cecal fermentation in rats fed a HAS diet (Experiment 2).
Food intake and body weight gain were lowest in rats fed PP (Table 3). Food intake was highest in rats fed RP or SP and intermediate in the casein-fed group. Body weight gain was highest in rats fed SP and casein but did not differ from that in rats fed RP. The weight of cecal tissue in these rats (Table 3) was approximately double that of rats fed LAS (Table 2). Cecal tissue weight was lowest in rats fed RP or PP and highest in those fed casein with intermediate weights in rats fed SP. The wet weight of cecal contents in these rats (Table 3) was substantially higher than in rats fed LAS (Table 2). The wet weight of cecal contents followed the same pattern as cecal tissue weights, i.e., lowest in rats fed RP or PP, highest in those fed casein, and intermediate in rats fed SP. pH values of cecal contents (Table 3) were substantially lower than those in rats fed the LAS (Table 2). Cecal pH was related inversely to the cecal tissue weight (r = 0.711, P < 0.0001) and contents (r = 0.849, P < 0.0001) and was lower in rats fed casein or SP than in those fed RP or PP.
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0.675, P < 0.0003). The butyrate pool was lowest in rats fed casein and highest in those fed PP.
Effects of the graded levels of PP on cecal fermentation in rats fed a HAS diet (Experiment 3). Food intake was lowest in rats fed 25% PP but did not differ significantly from the 0 and 1% PP groups (Table 4). Intakes in the other groups did not differ from those in rats fed 0 and 1% PP. Body weight gain was lowest in rats fed 25% PP; the other groups did not differ from one another.
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Effects of the graded levels of PP on fecal excretions of nitrogen and starch in ileorectostomized rats fed a HAS diet (Experiment 4).
When ileorectostomized rats were fed diets containing PP substituted for casein in the range from 0 to 25% of the diet, the ratio of starch to nitrogen in feces decreased (Fig. 2). The ratio was 18.5 in rats fed 0% PP and 8.0 in those fed 25% PP. This difference was due solely to greater excretion of protein; that of starch was unaffected (data not shown).
Effects of an artificial resistant protein on cecal fermentation in rats fed a HAS diet (Experiment 5).
Food intake in rats fed HAS with casein or with artificial resistant protein was significantly lower than that in rats fed LAS with casein (Table 5). Despite this difference, body weight gain did not differ among groups. As observed in Experiments 1 and 2, cecal tissue weight, as well as the amount of cecal contents, in rats fed HAS with casein was significantly greater than that in rats fed LAS with casein. Values in rats fed HAS with artificial resistant protein did not differ from those in rats fed HAS with casein. The pH of cecal contents was lowest in rats fed HAS with casein, intermediate in those fed HAS with artificial resistant protein and highest in rats fed LAS with casein.
We believe that this is the first time that the effects of dietary protein and their interaction with starch types on large bowel SCFA have been examined in rats. As expected from previous studies with HAS in rats (Mallett et al. 1988
Manuscript received 20 October 1997. Initial reviews completed 12 November 1997. Revision accepted 18 February 1998.
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Fig 1.
Cecal pool-size (A) and concentration (B) of butyrate and succinate in rats fed 0, 1, 3, 5, 10 and 25% potato protein diets containing 200 g/kg of high amylose cornstarch for 14 d (Experiment 3). Analyzed feces were collected for the last 3 d of the experimental period. Each point is expressed as mean ± SEM (n = 6); points for an organic acid with different superscript letters are significantly different (P < 0.05).
0.345, P < 0.0394). Fecal nitrogen excretion rose with increasing dietary PP (r = 0.775, P < 0.0001). Fecal neutral sterols and coprostanol/cholesterol ratios (mol/mol) increased with increasing dietary PP level, and fecal neutral sterol excretions in rats fed the 5, 10 and 25% PP were significantly higher than those in rats fed the 0% PP. Fecal coprostanol/cholesterol ratios in rats fed the 3, 10 and 25% PP diets were also significantly higher than those in rats fed 0% PP (Table 4).
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Fig 2.
Fecal starch/nitrogen ratio in ileorectostomized rats fed 0, 1, 3, 5, 10 and 25% potato protein diets containing 200 g/kg of high amylose cornstarch for 10 d (Experiment 4). Each point is expressed as mean ± SEM (n = 6); points with different superscript letters are significantly different (P < 0.05).
View this table:
Table 5.
Effects of an artificial resistant protein on food intake, body weight gain, cecal weight, cecal organic acids and fecal excretion in rats fed a high amylose cornstarch diet for 10 d (Experiment 5)1
DISCUSSION
Abstract
Introduction
Methods
Results
Discussion
References
) and in pigs (Brown et al. 1997), we found that its consumption resulted in significantly higher large bowel pools of SCFA compared with LAS (Tables 2 and 3). These higher SCFA levels are consistent with the greater RS content of the former, leading to a greater provision of fermentative substrate for the large bowel microflora. This hypothesis is supported in these experiments by the greater weight of cecal contents (including starch) in rats fed HAS in comparison with those fed LAS (Tables 2 and 3). In earlier studies (Brown et al. 1997, Mallett et al. 1988) with HAS, data supporting greater large bowel fermentation were reported principally for acetate, propionate and butyrate and minor SCFA. Other, potentially important, carboxylic acids were not recorded. However, the present data show clearly that, even in rats fed LAS, cecal succinate was present in substantial amounts and (depending on protein source in diet) equaled or exceeded butyrate (Table 2). In rats fed HAS with casein, the most predominant cecal carboxylic acid was not a SCFA, but succinate, whose cecal pool size reached almost 600 µmol (Table 3). This greater pool of succinate could be a simple reflection of greater fermentation. However, this cannot be the sole reason; in rats fed HAS with casein, succinate level was comparable to that of acetate, but when RP, PP or SP were fed instead of casein, the contribution of succinate was much less, whereas that of butyrate was higher (Table 3). In rats fed the HAS diet, the most striking succinate-lowering effect was found with RP, whereas PP was most effective in increasing butyrate (Table 3). Thus, the protein source had a substantial influence on the fermentation products of HAS, possibly due to the different digestibilities of each protein. The fecal coprostanol/cholesterol ratios were higher in rats fed HAS with RP, PP or SP than in rats fed HAS with casein (Table 3), suggesting that RP, PP or SP may alter the relative proportions of the cecal bacterial species through the supply of nitrogen (undigested proteins) to the cecum. Therefore, the differences in fermentation profiles of HAS among rats fed RP, PP or SP and casein may have been due to differences in the cecal microflora.
). However, in this experiment, the smaller succinate pool in rats fed HAS with RP or PP was not accompanied by any change in the pool of propionate, and the only consistent change was in butyrate (Table 3). Nevertheless, the greater pool of succinate in rats fed HAS relative to those fed LAS has implications for large bowel physiology. The most obvious consideration is pH. Cecal pH in rats fed HAS was considerably lower than that in rats fed LAS (Tables 2 and 3). In rats fed HAS with casein, RP, PP or SP, there was an inverse relationship between cecal pH and succinate pool size (r = 0.675, P < 0.0003) (Table 3). Although the precise mechanism for succinate absorption in the cecum and colon is largely unknown, succinate is likely to be absorbed slowly, as is lactate (Hoshi et al. 1994). On the basis of this premise, the substantially lower cecal pH in rats fed diets that raised succinate pools (compared with similar diets which did not) would be due primarily to the accumulation of this acid.
, Lupton and Kurtz 1993, Sakata 1987). However, data from this study in rats fed HAS show that the greater SCFA pools cannot be the sole reason for the greater cecal tissue weight because in rats fed HAS with casein, cecal tissue weights were higher than in those fed HAS with RP or PP despite the similar amounts of cecal total SCFA (acetate, propionate and butyrate) (Table 3). A more probable explanation is that the greater pool size of succinate is likely to play a role in stimulating the growth of cecal tissue, either by itself or by lowering cecal pH as previously described in rats fed certain oligosaccharides (Hoshi et al. 1994). This hypothesis is also supported by the facts that cecal tissue weight was inversely related to cecal pH and was positively related to cecal succinate (Table 3).
, Mason 1984). Both possibilities may have occurred. However, the precise contribution of undigested protein and increased bacterial protein to fecal nitrogen excretion remains to be established.
, MacFarlane et al. 1986, Mason 1984). However, when a highly purified and digestible protein such as casein is the sole source in the presence of large amounts of fermentable carbohydrates such as HAS, nitrogen supply might become insufficient to sustain a rapid bacterial proliferation. Studies in rats fed high levels of RS have shown that the flux of ammonia into the cecum was increased greatly (Rémésy and Demigné 1989). If the demands of fermentation were too great, nitrogen deficiency could result. This may be exacerbated by the fact that the rate of fermentation of HAS may be relatively rapid (Topping et al. 1997). Thus, one possibility is that an imbalance may have occurred in the ratio of carbohydrate and nitrogen in rats fed HAS with casein. In this connection, the starch/nitrogen ratio of the ileal effluent in rats fed HAS with 0-25% PP drastically changed as a function of dietary PP level (Fig. 2), suggesting that feeding PP should correct the imbalance in the ratio of carbohydrate/nitrogen in rats fed HAS with casein. Other proteins with lower digestibility (i.e., RP and SP) may also correct the imbalance. In a separate study, in which rats were fed a commercial diet supplemented with HAS (200 g/kg diet), almost all of the cecal carboxylic acids were SCFA, and succinate concentrations were almost undetectable (Morita, Kasaoka and Kiriyama, unpublished observations). Commercial diet contains more than 4% crude fiber and 20-30% protein. Some of the protein may be trapped in the plant cell matrix and escape complete digestion in the small intestine. In addition, some poorly fermentable fibers (e.g., lignocellulose) in a commercial diet may allow the bacterial population to increase by providing them with attaching surfaces. Therefore, it appears reasonable to hypothesize that accumulation of cecal succinate originated from an imbalance in the carbohydrate/nitrogen ratio, which affected the cecal microflora. This hypothesis was reinforced by the facts that the replacement of casein by 6% artificial resistant protein in the HAS diet significantly lowered cecal succinate, increased pH, and improved the productivity of SCFA as well as cecal fermentability of HAS compared with those variables in rats fed the HAS diet with casein (Table 5). Thus, resistant proteins and possibly resistant peptides play an important role in correcting an imbalance in the ratio of carbohydrate to nitrogen and controlling the cecal fermentation of RS.
and Chacko and Cummings (1988) have reported that up to 12 g of protein may escape digestion and enter the human colon in people consuming a Western diet. A meta-analysis of dietary studies has shown that greater intake of protein is associated with enhanced risk of colon cancer (Cassidy et al. 1994). This may reflect greater accumulation of potentially toxic by-products of protein metabolism such as phenol, p-cresol, indoles, amines and ammonia in the colon (MacFarlane and Cummings 1991). Clearly, these data are in contrast to the current experiments in rats, and care must be taken when we refer to the balance of RS and resistant protein in the human diet. Most studies evaluating the potential benefits of RS to produce SCFA, however, have been conducted in rat models consuming purified or semipurified diets that contain highly digestible protein and a relatively large amount of RS (usually 10-30% in diets). Thus, caution must be exercised in designing such experiments, especially in the sources of protein that are used. To date, only digestibility and amino acid composition have been recognized as important factors for the evaluation of quality of dietary proteins (FAO/WHO 1990). However, it appears possible that resistant proteins and peptides may have an important physiologic role through their interaction with RS in providing butyrate to colonic tissues.
1
Presented in part at The Research Committee of Essential Amino Acids, March 22, 1996, Tokyo, Japan. Specific effects of resistant protein on cecal fermentation in rats. Research Committee of Essential Amino Acids (Japan), p. 19. Abs. 146.
FOOTNOTES
2
The costs of publication of this article were defrayed in part
by the payment of page charges. This article must therefore be hereby marked "advertisement"
in accordance with 18 USC section 1734 solely to indicate this fact.
3
To whom correspondence should be addressed.
4
Abbreviations used: HAS, high amylose cornstarch; LAS, low amylose cornstarch; PP, potato protein; RP, rice protein; RS, resistant starch; SCFA, short-chain fatty acids; SP, soybean protein.
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ACKNOWLEDGMENT |
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We thank T. Hisatsune (UnicoopJapan) for transmitting the information on potato protein production.
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LITERATURE CITED |
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Abstract
Introduction
Methods
Results
Discussion
References
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Z. Liu, W. Ishikawa, X. Huang, H. Tomotake, J. Kayashita, H. Watanabe, and N. Kato A Buckwheat Protein Product Suppresses 1,2-Dimethylhydrazine-Induced Colon Carcinogenesis in Rats by Reducing Cell Proliferation J. Nutr., June 1, 2001; 131(6): 1850 - 1853. [Abstract] [Full Text]
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Z. Guo, C. Yuan, T. Wei-Lavery, Y. Fang, R. A. Garvin, H. I. Nishida, and T. Nishida Secretion of Phospholipid Transfer Protein by Human Hepatoma Cell Line, Hep G2, Is Enhanced by Sodium Butyrate J. Nutr., November 1, 1999; 129(11): 1984 - 1991. [Abstract] [Full Text]
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T. Morita, S. Kasaoka, K. Hase, and S. Kiriyama Psyllium Shifts the Fermentation Site of High-Amylose Cornstarch toward the Distal Colon and Increases Fecal Butyrate Concentration in Rats J. Nutr., November 1, 1999; 129(11): 2081 - 2087. [Abstract] [Full Text]
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T. Morita, S. Kasaoka, K. Hase, and S. Kiriyama Oligo-L-Methionine and Resistant Protein Promote Cecal Butyrate Production in Rats Fed Resistant Starch and Fructooligosaccharide J. Nutr., July 1, 1999; 129(7): 1333 - 1339. [Abstract] [Full Text]
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