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Article

Rye Bread Improves Bowel Function and Decreases the Concentrations of Some Compounds That Are Putative Colon Cancer Risk Markers in Middle-Aged Women and Men^{1 ,2}

Soile M. Gråsten³, Katri S. Juntunen, Kaisa S. Poutanen^*, Helena K. Gylling, Tatu A. Miettinen and Hannu M. Mykkänen

University of Kuopio, Department of Clinical Nutrition, FIN-70211 Kuopio, Finland; ^* VTT Biotechnology, FIN-02044 VTT, Finland; and University of Helsinki, Department of Medicine, Division of Internal Medicine, FIN-00029 HYKS, Helsinki, Finland

³To whom correspondence and reprint requests should be addressed.

	ABSTRACT

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Cereal fiber may reduce the risk of colorectal cancer by diluting colonic contents due to increased fecal output, by accelerating intestinal transit, by increasing fecal frequency and by altering bacterial metabolism. The effects of whole-meal rye bread on some putative colon cancer risk markers were investigated in 17 healthy Finnish subjects using a randomized crossover trial with two 4-wk bread consumption periods and a 4-wk washout period between the bread periods. White wheat bread was used as a control. Test breads covered a minimum of 20% of the daily energy intake (range, 4330–14,033 kJ/d). Intestinal transit time, stool weight, fecal bacterial enzyme activities and short-chain fatty acid, ammonia, diacylglycerol (DAG) and bile acid concentrations in feces (expressed per gram wet feces) were measured. Whole-meal rye bread significantly increased fecal output and fecal frequency and shortened mean intestinal transit time compared with wheat bread in both women and men. Activities of ß-glucuronidase and ß-glucosidase (expressed per gram wet feces) were significantly lower in men and urease activity significantly higher in women during the rye bread period (RBP). Fecal butyrate concentration was higher during the RBP in men. Fecal ammonia and DAG concentrations did not differ between bread periods. Fecal total and secondary bile acid concentrations were significantly lower during RBP in both women and men. This study shows that whole-meal rye bread significantly improves bowel function in healthy adults and may decrease the concentration of some compounds that are putative colon cancer risk markers.

KEY WORDS: • humans • rye bread • wheat bread • bowel function • short-chain fatty acids • bacterial enzymes • bile acids

	INTRODUCTION

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A diet rich in high fiber cereal may be associated with a reduced risk of colorectal cancer (ECP Consensus Panel 1998). Evidence from both epidemiologic and case control studies suggests that cereal fiber may protect the colon from cancer development (Hill 1997 , Jacobs et al. 1998 ), although recent prospective studies failed to show any protective effect by cereal fiber (Fuchs et al. 1999 , Platz et al. 1997 ). Physiologic effects of dietary fiber in the large intestine depend on the fermentability of the fiber, which is influenced by chemical composition, solubility, physical form and the presence of lignin and other compounds (Stephen 1994 ).

In grains, insoluble lignified fiber in the outer bran is degraded only to a small extent and increases fecal bulk by its physicochemical properties and water-holding capacity (Bach Knudsen et al. 1997 ). The larger bulk dilutes carcinogens, mutagens and tumor promoters, resulting in a lower risk of colon cancer (Weisburger et al. 1993 ). Larger bulk is associated with decreased intestinal transit time (Cummings et al. 1992 ), reducing the contact time of colonic epithelial cells with carcinogens or tumor promoters. On the other hand, soluble fibers in grains are fermented readily by the colonic bacteria, exerting only marginal effects on fecal weight and intestinal transit time (Bach Knudsen et al. 1997 ). However, soluble fibers may increase stool mass by increasing bacterial cell mass (Stephen and Cummings 1980 ). These fibers also have a potential to modify the metabolism of colon carcinogens, yielding detoxified products and thereby possibly reducing colon carcinogenesis (Weisburger et al. 1993 ); some soluble fibers, however, may also enhance cancer development (Harris and Ferguson 1999 ). The short-chain fatty acids (SCFA),⁴ acetate, propionate and butyrate, are physiologically important end products of colonic fermentation. Butyrate is the preferred fuel for colonocytes, especially in the distal colon and may be a protective factor in colon carcinogenesis (Scheppach 1998 ). SCFA may also decrease the colonic pH and inhibit dehydroxylation of bile acids, thus inhibiting conversion of primary bile acids to secondary bile acids (Christl et al. 1997 )

Bacterial fermentation in the colon produces large amounts of end products, some of which have been shown to be harmful to the colonic epithelium and are putative colon cancer risk markers. Bacterial ß-glucosidase hydrolyzes plant glycosides to release aglygones, many of which are mutagenic, although some also have anticarcinogenic activity (Rowland 1995 ). Bacterial ß-glucuronidase may form toxic compounds by releasing aglygones from glucuronide conjugates formed in the liver. Bacterial urease produces ammonia from urea. Ammonia is considered to be a potential tumor promoter in the colon and has been postulated to enhance neoplastic transformation in the gut (Clinton et al. 1988 ). Intestinal bacteria convert primary bile acids to secondary bile acids, which are also thought to promote the tumorigenic process in colon cancer (Narisawa et al. 1974 , Reddy et al. 1976 , Reddy and Watanabe 1979 ). A high ratio of lithocholic acid (LCA) to deoxycholic acid (DCA) is proposed to increase colon cancer risk (Owen at al 1986 ). Intestinal bacteria are also capable of producing diacylglycerols (DAG) from phospholipids and dietary fat (Morotomi 1990 ); this activity is enhanced by bile acids, especially DCA and chenodeoxycholic acid (CDCA). Diacylglycerols are important activators of protein kinase C (PKC) isozymes (Nishizuka 1992 ). Different PKC isozymes stimulate cell proliferation (Chapkin et al. 1993 ); thus, increased DAG concentration may cause a chronic state of increased cell proliferation.

The amount and type of substrate available to the microflora are important factors controlling bacterial metabolism in the colon. These substrates may affect the end products of bacterial metabolism contributing to the colon cancer risk. The modifying effect of dietary fiber on some putative colon cancer risk markers depends on the type of fiber consumed (Reddy 1999 ). Rye is a traditionally used fiber-rich cereal in Finland and other Northern countries. It is a rich source of insoluble and soluble arabinoxylans; it also contains >2% indigestible fructan (Åman et al. 1997 , Härkönen et al. 1997 ). In Finland, 40% of the dietary fiber intake originates from rye (National Public Health Institute 1998 ).

In this study, we determined the effects of whole-meal rye bread on bowel function and previously mentioned putative colon cancer risk markers. It was hypothesized that replacing customarily consumed cereal products with fiber-rich whole-meal rye bread would influence bowel function and the metabolic activity of the intestinal flora in a favorable manner, compared with white wheat bread containing low amounts of cereal fiber.

	MATERIALS AND METHODS

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Subjects.

Healthy Finnish volunteers from the Kuopio area (n = 17; 9 women and 8 men) participated in the study. The ages of the women and men [mean ± SD (range)] were 40.6 ± 7.7 (28–51) and 43.4 ± 9.9 (31–56) y; weights were 64.7 ± 7.7 (53.1–76.4) and 86.0 ± 7.4 (76.4–99.2) kg; and body mass indices were 22.9 ± 2.5 (20.0–28.6) and 27.8 ± 2.0 (25.0–30.3) kg/m², respectively. Each subject gave a written informed consent before participation in the study, which was approved by the Ethics Committee of Kuopio University Hospital.

Study design.

The study was a randomized crossover trial. The first bread period was preceded by a 2-wk baseline period. At the beginning of the study, the subjects were advised to maintain their body weight and lifestyle habits (exercise, alcohol consumption, smoking) unchanged and not to use foods that affect bowel function (plums and plum juice, dried fruits, brans, muesli, various seeds and licorice). The subjects kept 4-d food records during the baseline period to determine individual energy intakes. After the baseline period, the subjects were divided randomly into two groups and advised to consume either rye breads or wheat breads instead of customarily used breads and baked products for 4 wk. After a 4-wk washout period, the groups were reversed. For premenopausal women, the test bread periods were started on d 5–10 from the beginning of menstruation and the length of the bread periods was adjusted to cycle length.

Diet.

The composition of the test diets was described previously (Leinonen et al. 2000 ). The subjects were advised to eat a minimum of 20% of their daily energy intake in the form of test breads. During the rye bread period (RBP), the customarily used breads and baked products were replaced by rye breads and during the wheat bread period (WBP), by wheat breads. In addition, the subjects were allowed to eat one piece of sweet pastry or a portion of porridge once a day. Pasta and rice products were allowed to be consumed as a part of warm dishes. Otherwise, the diet was to be constant. In particular, the subjects were advised not to change their consumption of fiber-containing foods such as vegetables, fruits and berries. The test breads (four commercially available rye breads and six wheat breads) were obtained from Fazer Bakeries Ltd, Lahti, Finland and Vaasan & Vaasan Ltd, Helsinki, Finland. Fresh-baked breads were available once a week from the study center.

The participants were given written instructions concerning the diet, and a clinical nutritionist advised them about practical management of the diet. If sliced breads were not available, the subjects were given detailed instructions how to slice the loaves, to guarantee that the subjects ate the correct amount of test breads. A minimum of 4–5 portions of test breads had to be eaten, and the number of test bread portions to be eaten varied according to the daily energy intake. The consumption of the test breads and other cereals, as well as fecal frequency and possible gastrointestinal side effects, was recorded daily and dietary adherence was followed by 4-d food records during the last 2 wk of both bread periods. A clinical nutritionist calculated nutrient intake using Micro-Nutrica, a software nutrient calculation program for nutrients (Social Insurance Institution, Helsinki, Finland and the database of Finnish foods; Rastas et al. 1993 ). The nutrient composition of the test breads was analyzed by VTT Biotechnology (Espoo, Finland) and added to the database.

Fecal sample collection and analysis.

All men and postmenopausal women collected stool specimens for 5 d during wk 4 of each test bread period. Due to adjustment to the menstrual cycle length, the premenopausal women collected stools in wk 3 of both test bread periods. The subjects could either bring stools daily to the study center or collect all feces for 5 d and bring all samples at the end of collection period. At home, the subjects were advised to store feces in a cold box (10°C) until they were transported to the study center. In the laboratory, the fecal samples were stored at -20°C until determination of the wet and dry weights, intestinal transit time, bacterial enzyme activities and SCFA, DAG, ammonia and bile acid concentrations.

For measurement of intestinal transit time, the subjects were administered Sitzmarks radiopaque markers (Konsyl Pharmaceuticals, Fort Worth, TX) on the first morning of each feces collection period and collected feces for five consecutive days. Fecal samples were X-rayed, and the mean transit time was calculated as a mean time for rings to pass through the intestinal tract as described by Corazziari et al. (1987) .

Frozen stools were weighed, thawed at 4°C and pooled by adding 20 g/100 g distilled water and homogenizing the mixture with a Stomacher laboratory blender before analyzing the enzyme activities and fecal metabolites. Activities of fecal bacterial enzymes were determined as described by Ling et al. (1994) . Briefly, pooled samples were thawed at 4°C and homogenized with 0.1 mol/L potassium phosphate buffer, pH 7.0 (1:9; wt/wt). The mixture was sonicated (3 x 20 s) in an ice bath, centrifuged at 500 x g for 15 min and the supernatant used for analysis. Activities of ß-glucuronidase (EC 3.2.1.31) [substrate phenolphtalein mono-ß-D-glucuronic acid (Sigma, St. Louis, MO) in 0.1 mol/L potassium phosphate buffer, pH 6.8], and ß-glucosidase (EC 3.2.1.21) [substrate p-nitrophenyl-ß-D-glucopyranoside (Sigma) in 0.1 mol/L potassium phosphate buffer, pH 7.4] were determined at 37°C as described by Freeman (1986) . Activity of fecal urease (EC 3.5.1.5) was determined by incubating the fecal supernatant fluid for 10 and 20 min in 0.02 mol/L potassium phosphate buffer, (pH 7.4) containing urea as substrate. The ammonia released in the reaction was determined using an ammonia electrode (model no. 95–12; Orion, Helsinki, Finland). To express enzyme activities as nmol substrate metabolized/(min·mg protein), protein in the fecal supernatant fluid was determined in duplicate using the method of Lowry et al. (1951) . Bovine serum albumin was used as a standard. Enzyme activities were also expressed as nmol substrate metabolized/(min·g wet feces). Fecal dry weight was determined by oven drying at 105°C for 17 h.

Fecal SCFA were measured by the method of Schooley et al. (1985) . Samples of the homogenized fecal samples (1 g) were suspended in 9 g/L NaCl, spiked with 500 µL internal standard (heptanoic acid, final concentration 1 mmol/L), and extracted twice with 2.5 mL diethyl ether. A standard mixture of pure SCFA was treated similarly. The combined ether extracts were analyzed by gas chromatography (HP 5890; Hewlett Packard, Wilmington, DE) using a DB-WAX column (30 m, 0.5-µm film). SCFA were separated over a temperature range of 50–230°C (8–12°C/min gradient) using helium as the carrier gas and detection with a flame ionization detector

Fecal DAG concentration was determined by a radioenzymatic assay using a commercial kit (Amersham International, Amersham, UK) based on the Escherichia coli DAG kinase method (Preiss et al. 1986 ). Fecal samples were thawed at 4°C and homogenized with potassium phosphate buffer, pH 7.0 (1:15 wt/wt). DAG were extracted from the homogenate as described by Phan et al. (1991) . For separation of [³²P]-phosphatidic acid, Amprep minicolumns (Amersham International) were used; the amount of [³²P]-phosphatidic acid was quantitated by liquid scintillation spectrometry (LKB Wallac 1215 rackbeta, Wallac, Turku, Finland).

Ammonia concentration was measured spectrophotometrically (Chaney and Marbach 1961 ). Samples were thawed at 4°C, mixed with 3 g/L trichloroacetic acid (1:10 wt/wt) (Lin and Wisek 1991 ) and centrifuged at 500 x g for 12 min; the ammonia concentration was determined from the supernatant fraction., with ammonium chloride as a standard.

Fecal bile acids were measured by GLC (Grundy et al. 1965, Miettinen 1982 ) on a 50-m long SE-30 capillary column (Hewlett Packard, Little Falls, Wilmington, DE).

Statistical analysis.

The SPSS/Win program (Chicago, IL) was used for statistical analysis of the data. Due to the small number of participants, differences between bread periods in all variables were tested with the nonparametric Wilcoxon signed-ranks test. All results are expressed as means ± SD The differences between periods were considered to be significant at P < 0.05.

	RESULTS

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Diet.

The subjects consumed the recommended amount of test bread portions during both test bread periods (Table 1 ). The men ate significantly more bread than the women during both bread periods (P < 0.01). The mean fiber intake from the test breads was significantly greater during RBP compared with WBP in both women (P < 0.01) and men (P < 0.05); the difference was 13.5 and 19.1 g for women and men, respectively (Table 2 ). Energy, fat, protein and carbohydrate intakes did not differ during the rye and wheat bread periods.

Fecal wet weight was significantly greater during RBP than WBP in both women and men (P < 0.05), but the percentage of dry matter in feces did not differ between the bread periods (Table 3). Mean intestinal transit time was significantly shorter during RBP in men (P < 0.05) and tended to be shorter (P = 0.07) in women. The difference in mean intestinal transit time between the bread periods was 11 h for women and 8 h for men. Fecal frequency was significantly greater during RBP in both women and men (P < 0.05).

Fecal bacterial ß-glucuronidase, ß-glucosidase and urease activities are presented in Table 4 . When enzyme activities were expressed as nmol/(min·mg protein), urease activity in women was higher during the rye bread period (P < 0.05), but there was no difference in ß-glucuronidase and ß-glucosidase activities between the bread periods. In men, ß-glucuronidase activity tended to be lower during RBP compared with WBP (P = 0.07). ß-Glucosidase and urease activities did not differ between bread periods in men. When enzyme activities were expressed as nmol/(min·g wet feces), activities of ß-glucuronidase and ß-glucosidase were significantly lower during RBP in men but not in women. Activity of urease was greater during RBP in women (P < 0.05) but not in men.

Total SCFA (in women 43.6 ± 2.2 vs. 39.3 ± 4.4 and in men 48.2 ± 3.8 vs. 45.0 ± 4.4 mmol/kg wet feces), acetate (in women 27.0 ± 1.2 vs. 26.3 ± 3.1 and in men 32.0 ± 2.7 vs. 29.4 ± 3.2 mmol/kg wet feces) and propionate (in women 5.8 ± 0.4 vs. 5.8 ± 0.7 and in men 7.0 ± 0.1 vs. 7.2 ± 0.9 mmol/kg wet feces) concentrations did not differ between the rye bread and wheat bread periods, respectively. Butyrate concentration was significantly higher in men during RBP (10.2 ± 1.1 vs. 7.5 ± 0.7 mmol/kg wet feces, P < 0.05), but there was no difference in butyrate concentration in women between the RBP and WPB (7.4 ± 0.6 vs. 6.1 ± 0.7 mmol/kg wet feces).

Fecal DAG concentrations did not differ between RBP and WBP (85 ± 34 vs. 84 ± 28 nmol/g wet feces in women and 87 ± 51 vs. 111 ± 44 nmol/g wet feces in men). One man had a very high fecal DAG concentration (472 and 114 nmol/g wet feces during RBP and WBP, respectively) and his results were not included in the statistical analysis of the data. The ammonia concentration did not differ between the rye bread and wheat bread periods in women (41.2 ± 19.4 vs. 40.4 ± 20.2 µmol/g wet feces) and in men (30.0 ± 10.4 vs. 35.3 ± 14.3 µmol/g wet feces).

Fecal total bile acid concentration was lower during the RBP (P < 0.05) in both women and men (Table 5 ). There were significant differences in proportions of some fecal bile acids from the total bile acid pool between RBP and WPB. The proportion of cholic acid (CA) was greater in women (P < 0.05) and that of CDCA was greater in men (P < 0.05) during the RBP. The proportions of secondary bile acids LCA and epideoxycholic acid were smaller in women (P < 0.05) and those of isolithocholic acid and epideoxycholic acid were smaller in men (P < 0.05) during RPB. Proportions of ursodeoxycholic acid in women (P < 0.05) and ursodeoxycholic acid and ketoacids in men (P < 0.05) were greater during RBB. The DCA/CA and LCA/CDCA ratios did not differ between the test bread periods when the data were analyzed separately for women and men. However, when the data for women and men were pooled, the difference between the bread periods was significant (P < 0.05). The LCA/DCA ratio was significantly lower during the rye bread period in women (P < 0.05) but not in men.

	DISCUSSION

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Bowel function (fecal weight, intestinal transit time and fecal frequency) was significantly improved during the rye bread consumption period in both men and women. It has been shown that intake of dietary fiber, especially insoluble fiber increases fecal bulk and the amount of stools passed. In this study, the intake of total and also insoluble dietary fiber was four- to fivefold during the rye bread period compared with the wheat bread period and the effects of rye bread consumption on bowel function were thus expected. In comparing populations having different fecal weights, low fecal weight is associated with increased risk of colon cancer (Cummings et al. 1992 ). The authors postulated that at stool weights of 100 g/d, colon cancer risk is high; when stool weights are > 200 g/d, colon cancer risk becomes low. In our study, the mean fecal weight exceeded 200 g/d during the rye bread period in both women and men.

It has been suggested that when stool weights exceed 150 g/d, only relatively small reductions in transit time are seen (Cummings 1992 ). However, in this study, the fecal weights varied from 151 (in women during the wheat bread period) to 335 g/d (in men during the rye bread period), and a significant decrease in transit times was seen during the rye bread period in both sexes.

The percentage of fecal dry matter was not affected, indicating that rye bread increased both dry and wet weights of feces. Intake of rye fiber may have affected fecal weight by several mechanisms. A large amount of insoluble fiber increases fecal bulk by its physicochemical properties and water-holding capacity. Rye bread also contains fermentable fiber, which can affect the bacterial metabolism and increase fecal weight by the formation of bacterial biomass. Faster transit also induces greater fecal bulk independently of the diet by increasing the formation of bacterial mass and excretion of nonstarch polysaccharides (Stephen et al. 1987 ).

Activities of fecal enzymes, expressed as nmol/(min·mg protein) differed significantly between the bread periods only for urease in women. These data do not support the hypothesis that rye bread alters the metabolic activity of the colonic flora in humans. However, the finding that ß-glucuronidase and ß-glucosidase activities expressed as nmol/(min·g wet feces) were lower in men during the rye bread period further confirms the dilution effect of the colonic contents. The difference in fecal wet weights of men between the test bread periods was greater than that of women (137 g vs. 52 g). Therefore, the dilution effect in ß-glucuronidase and ß-glucosidase activities was not observed in women. Both ß-glucuronidase and ß-glucosidase have been implicated in the generation of mutagens or carcinogens; ß-glucuronidase, in particular, seems to be important in colon carcinogenesis due to its wide substrate specificity (Goldin 1990 ). Lower activities of these enzymes expressed as nmol/(min·g wet feces) can be considered beneficial in terms of the risk of colon cancer. There are no previous studies on the effects of whole-meal rye on bacterial metabolic activity in humans. Ling et al. (1994) investigated the effect of a rye fiber product, consisting mainly of the aleurone layer and a probiotic bacterial strain, on fecal bacterial enzymes in healthy women. They reported that rye fiber alone did not affect ß-glucuronidase or ß-glucosidase activities but did lower urease activity. In our study, the rye bread consumed also provided polysaccharides other than those in the aleurone layer of rye kernel. The reason for higher urease activity in women during the rye bread period in the present study is unclear. Earlier, it was shown in rats that rye bran enhances the growth of bifidobacteria (Ryhänen et al. 1996 ), and bifidobacteria possess a low ß-glucuronidase activity (Hawksworth et al. 1971 ). In our study, there was a trend toward lower activity of ß-glucuronidase expressed as nmol/(min·mg protein) in men during the rye bread period. It is possible that, due to the insufficient statistical power of the study, our data could not support the hypothesis that whole-meal rye alters the metabolic activity of the intestinal flora.

Fecal butyrate concentration was higher during the rye bread period in men, although total SCFA concentrations were similar. All SCFA are important substrates for colonocytes; of these SCFA, butyrate is the most important, especially in the distal colon (Scheppach 1998 ). Butyrate may be a putative protective factor in colon carcinogenesis; its role is not yet clear, however, because it has different effects in normal and neoplastic cells (Scheppach 1998 ). In vitro data have shown butyrate to promote differentiation and apoptosis (Deng et al. 1992 , Hague et al. 1996 ), but in vivo butyrate has been shown to stimulate cell proliferation (Lupton and Kurtz 1993 ). Because fecal bulk was much greater during RBP, the amount of total SCFA and butyrate produced by the fermentation also increased compared with WBP. A difference in the fecal concentration of butyrate between bread periods indicates a difference in colonic metabolism between bread periods.

Fecal DAG concentrations did not differ during the rye and wheat bread periods. Lack of the dilution effect in DAG concentration indicates a greater DAG production during the rye bread period. Increased intake of rye fiber during the rye bread period may have increased the amount of fat entering the colon (Zhang et al. 1994 ), and this fat served as a substrate for DAG production. High fiber intake might also have increased the metabolic activity or amount of DAG-producing bacteria. However, we did not observe a change in fecal enzyme activity expressed as nmol/(min·mg protein). Reddy et al. (1994) reported that dietary wheat bran, but not oat or corn bran, decreased the concentration of total DAG compared with a low fiber control diet. However, wheat bran also increased the amount of fecal fat excreted. The authors suggested that the type of dietary fiber and type of fat consumed may affect the amount of DAG produced in the gut.

Ammonia concentrations were similar between the test bread periods. In men, there was a trend toward lower concentration, but this difference was not significant. A similar trend was seen also in urease activity in men. In women, urease activity was greater during the rye bread period, but there was no similar difference in ammonia concentration between the test bread periods. Ammonia concentrations in our samples were markedly higher compared with previous studies (Birkett et al. 1996 and 1997 ). This may be due to differences between the studies in sample preparation.

Total bile acid concentration was lower during the rye bread period due to lower secondary bile acid concentrations. The lower concentration was likely due mainly to increased fecal bulk during the rye bread period. Yet, small differences in the bile acid profiles between the test bread periods and a trend toward smaller secondary to primary bile acid ratios may indicate that the increased load of fermentable fiber also altered the metabolism of bile acids in the colon. It has been shown that rye bread decreases the concentration of free secondary bile acids by changing the mode of their conjugation (Korpela et al. 1992 ), which indicates altered metabolic activity of the intestinal flora. In this study, we determined only the total bile acid concentration and did not differentiate the bile acids in the saponifiable, conjugated and free bile acid fractions. Lower concentration of bile acids and especially secondary bile acids is considered beneficial in terms of colon cancer risk. In addition, a lower LCA/DCA ratio may indicate a reduced risk of colon cancer (Owen et al. 1986 ). In this study, rye bread increased the proportion of ursodeoxycholic acid in feces. Ursodeoxycholic acid in pharmacologic doses has been shown to have chemopreventive effects in the colon in animal studies (Earnest et al. 1994 , Invernizzi et al. 1997 ) and it inhibits cell proliferation in vitro (Martinez et al. 1998 ). Still, the effects of physiologic concentrations of ursodeoxycholic acid in the colon are unknown.

In conclusion, this study shows that consumption of a normal amount of whole-meal rye bread as a part of a habitual diet has favorable effects in the colon in terms of colon cancer risk. Rye bread improves bowel function by increasing fecal weight and fecal frequency and by shortening intestinal transit time. The favorable effects of rye bread observed in this study are explained mainly by the dilution of the colonic contents. Increased fecal weight lowers the concentration of fecal bile acids in women and men and decreases the concentration of enzymes that produce compounds that may adversely affect the colon epithelium in men. The effects of rye bread on bacterial enzyme activities and on compounds that are putatively related to colon cancer risk must be evaluated in studies with a larger study population.

	FOOTNOTES

 
¹ Presented in poster form at Functional Food Research in Europe 3rd Workshop, FAIR CT96–1028, October 1–2, 1998, Probdemo, Haikko Manor, Finland, and the 15-year Anniversary Symposium of the Department of Clinical Nutrition, University of Kuopio, March 25–26, 1999, Kuopio, Finland [Maijala, S., Leinonen, K., Poutanen, K. & Mykkänen, H. (1998) Effects of whole-meal rye bread on bowel function and fecal bacterial enzymes in healthy adults (abs.). Abstracts are published in VTT symposium 187 publications and in Kuopio University Publications D. Medical Sciences 171] and at International Conference on Diet and Prevention of Cancer with Special Emphasis on Chemoprevention of Cancer, May 28-June 2, 1999, Tampere, Finland [Maijala, S., Leinonen, K., Gylling, H., Miettinen, T. A. & Mykkänen, H. (1999) Rye bread decreases fecal concentrations of total and secondary bile acids (abs.).]

² Supported by the Fazer Bakeries Ltd, Vaasan & Vaasan Ltd and the Technology Development Center of Finland. Bakeries supplied the test breads used in the study.

⁴ Abbreviations used: CA, cholic acid; CDCA, chenodeoxycholic acid; DAG, diacylglycerol; DCA, deoxycholic acid; LCA, lithocholic acid; PKC, protein kinase C; RBP, rye bread period; SCFA, short-chain fatty acids; WBP, wheat bread period.

Manuscript received November 29, 1999. Initial review completed February 20, 2000. Revision accepted May 5, 2000.

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