American Journal of Clinical Nutrition, Vol. 69, No. 1, 135-139,
January 1999
© 1999 American Society for Clinical Nutrition
Original Research Communications |
Gas production in humans ingesting a soybean flour derived from beans naturally low in oligosaccharides1,2,3
Fabrizis L Suarez,
John Springfield,
Julie K Furne,
Troy T Lohrmann,
Phillip S Kerr and
Michael D Levitt
 |
ABSTRACT
|
|---|
Background: Ingestion of soy products may cause excessive intestinal gas. This gas results from colonic bacterial fermentation of the indigestible oligosaccharides raffinose and stachyose, which are present in high concentrations in legumes.
Objective: The objective of the study was to compare gas production and gaseous symptoms in healthy volunteers after ingestion of 34 and 80 g soy flour made from either conventional soybeans or soybeans naturally low in indigestible oligosaccharides.
Design: In a double-blind, randomized, crossover protocol, breath hydrogen (an indicator of carbohydrate malabsorption), flatus frequency, and abdominal symptoms were assessed after subjects ingested the soy products and after 2 control meals (rice or lactose-hydrolyzed milk).
Results: The sum of breath-hydrogen concentrations for 8 h was significantly greater (P < 0.005) after 34 g conventional soy (60.4 ± 9.4 ppm) than after low-oligosaccharide soy (34.3 ± 8.1 ppm). Greater differences were observed with 80-g doses: 157.9 ± 19.4 ppm after conventional soy and 50.8 ± 6.8 ppm after low-oligosaccharide soy (P < 0.001). Flatus frequency (7.5 ± 1.9 times/12 h) was significantly greater (P = 0.039) after ingestion of 80 g conventional soy than after the control, rice meal (3.2 ± 0.8 times/12 h), whereas flatus frequency after the low-oligosaccharide soy meal (3.9 ± 0.7 times/12 h) was comparable with that after the rice meal. There were no significant differences in the severity of other abdominal symptoms.
Conclusion: Soy flour derived from low-oligosaccharide soybeans resulted in less gas production than that derived from conventional soybeans.
Key Words: Soy flour • low-oligosaccharide soybeans • breath-hydrogen concentration • gaseous symptoms • humans • raffinose • stachyose
|
INTRODUCTION
|
|---|
Legumes characteristically contain high concentrations of the indigestible oligosaccharides raffinose [ß-D-fructofuranosyl-O-
-D-galactopyranosyl-(1
6)--D-glucopyranoside] and stachyose [ß-D-fructofuranosyl-O--D-galactopyranosyl-(16)--D-galactopyranoside-(16)--D-glucopyranoside]. The indigestibility of these soluble sugars results in their delivery to the colon, where they are rapidly fermented by the colonic flora, resulting in the production of copious gas (1). This gas production may play a role in the acceptability of soy products as a major food source for humans (1–4) and animals (5).
The final steps in the production of raffinose and stachyose in soybeans are catalyzed by the enzymes galactinol–sucrose galactosyltransferase (EC 2.4.1.82, raffinose synthase) and galactinol–raffinose galactosyltransferase (EC 2.4.1.67, stachyose synthetase), respectively (Figure 1
). Screening of a large number of soy seeds for this enzyme activity resulted in the identification of a seed with very low activity, and this seed was shown to have a low concentration of raffinose and stachyose. The studies described in this report were designed to compare gas production and gaseous symptoms in healthy human volunteers ingesting concentrates derived from conventional and low-oligosaccharide soybeans.
View larger version (19K):
[in this window]
[in a new window]
|
FIGURE 1. Metabolic pathway for the synthesis of raffinose and stachyose from sucrose in soybeans. Galactinol provides galactoses for the conversion of sucrose to raffinose and of raffinose to stachyose. UDP-Glu, uridine diphosphate glucose; UDP-Gal, uridine diphosphate galactose; Glc6P, glucose-6-phosphate.
|
|
|
SUBJECTS AND METHODS
|
|---|
Subjects
The soy products were evaluated in 2 double-blind, randomized, crossover studies, each involving 20 healthy volunteers. A total of 28 subjects (11 men and 17 women) aged 29–62 y were studied; 12 individuals took part in both studies. Exclusion criteria included the existence of intercurrent illness, antibiotic ingestion within the previous 2 mo, and allergy to soy products or aspartame. The studies were approved by the Human Subjects Committee of the Institutional Review Board at the Minneapolis Veterans' Affairs Medical Center, and informed consent was obtained from all subjects.
Protocols
Study 1
In random order, 20 subjects ingested each of 3 test meals separated by 1-wk washout periods: 1) 500 mL soymilk made from 34 g conventional soy flour in 500 mL water, 2) 500 mL soymilk made from 34 g low-oligosaccharide soy (DuPont Agricultural Products, Wilmington, DE) in 500 mL water, and 3) 500 mL lactose-hydrolyzed, 1%-fat cow milk as a negative control. The composition of the conventional and low-oligosaccharide soy flours are shown in Table 1. Lactose was hydrolyzed by the addition of 1.07 g of a lactase preparation made from Kluyveromyces lactis (Lactaid Inc, Pleasantville, NJ) to 1 L milk, which was then incubated for 48 h at 4 °C (6). An assay using an enzymatic method (Lactose/D-galactose test kit; Boehringer Mannheim Biochemical Inc, Indianapolis) showed negligible lactose at the end of the incubation period. Aspartame (820 mg Equal/L; NutraSweet Co, Deerfield, IL) was added to the 2 soy products to yield a sweetness roughly equivalent to that of the lactose-hydrolyzed milk. All products were ingested with a straw from sealed cardboard containers (Trio Supplies, Minneapolis) to obscure differences in the color of the drinks (6). The taste of the 2 soy products was indistinguishable; however, the lactose-hydrolyzed milk differed in taste from that of the soy products.
Study 2
Twenty subjects were randomly assigned to receive each of 3 test meals over a 3-wk period with a 1-wk washout period between tests: 1) pudding containing 80 g conventional soy flour, 2) pudding containing 80 g low-oligosaccharide flour, and 3) a negative control, 80 g white rice cereal (Kellogg, Battle Creek, MI). (Preliminary studies showed that ingestion of the rice pudding resulted in minimal breath-hydrogen production.) The puddings were produced by mixing the flours with 240 mL water, 50 drops of orange flavoring (McCormick & Co, Inc, Hunt Valley, MD), and 2 g aspartame. Once again, the soy puddings were indistinguishable in color and taste, whereas the rice pudding was distinguishable from the soy products.
To minimize baseline breath-hydrogen excretion on the day of the study, the subjects ingested a rice and hamburger dinner the night before the challenge (7) and then fasted until the next morning. At 
0600, the test meals were ingested as rapidly as possible (within 30 min). No food was allowed for the next 8 h, but water and black coffee were allowed ad libitum. After the 8-h test period, subjects were allowed to resume their regular diets. End-alveolar breath samples for measurements of hydrogen and methane concentrations were collected before ingestion of the test meals and hourly for the next 8 h. The severity of each abdominal symptom (fullness, nausea, bloating, abdominal pain, and subjective impression of rectal gas) was rated by the subject on a linear 6-cm scale (0 cm = none, > 0–1.5 cm = trivial, 1.5–3 cm = mild, 3–4.5 cm = moderate, and 4.5–6 cm = severe) (6). Symptom severity was recorded over 2 time periods: from the time of test meal ingestion until 1200 and from 1200 until dinner. Each flatus passage from the time of test meal ingestion until 1800 was recorded and arbitrarily rated as "small" or "large."
Analysis of breath gases
Hydrogen and methane were quantified by gas chromatography and carbon dioxide was quantified by infrared spectroscopy as described previously (8). In rare cases, samples had a carbon dioxide concentration < 3.75% and were thus considered to have excessive atmospheric contamination; such a value was assumed to be the mean of the concentration obtained 1 h before and 1 h after the contaminated sample.
Statistical analysis
Data were analyzed by repeated-measures analysis of variance (ANOVA) (9, 10). The differences between treatments were calculated by paired t test, and the P value was adjusted by Bonferroni correction. A binomial distribution was used to calculate the P value (two-tailed). The sum of breath-hydrogen concentrations and mean symptom scores for each treatment period were compared.
|
RESULTS
|
|---|
Study 1
Breath-hydrogen concentrations resulting from the ingestion of 34 g of the 2 soymilks and the lactose-hydrolyzed cow milk are shown in Figure 2. The sum of breath-hydrogen concentrations from 1 to 8 h after ingestion of lactose-hydrolyzed milk (39.5 ± 9.4 ppm) and low-oligosaccharide soymilk (34.3 ± 8.1 ppm) were not significantly different, whereas breath-hydrogen excretion after ingestion of conventional soymilk (60.4 ± 9.4 ppm) was significantly greater than that after the other 2 treatments (overall P = 0.005; P = 0.006 when compared with low-oligosaccharide soymilk; and P = 0.033 when compared with lactose-hydrolyzed milk). Neither the severity of symptoms nor flatus frequency differed significantly with either soy product from that observed with the control, lactose-hydrolyzed milk (data not shown).
Study 2
Breath-hydrogen concentrations after ingestion of puddings containing 80 g of the soy concentrates or rice are shown in Figure 3. Although there appeared to be a trend toward greater total breath-hydrogen concentrations after the low-oligosaccharide soy (50.8 ± 6.8 ppm) than after the rice control (28.5 ± 6.1 ppm), this difference was not significant by ANOVA (P = 0.325). Total breath-hydrogen concentrations after conventional soy (157.9 ± 19.4 ppm) were significantly greater (P < 0.001) than concentrations observed after each of the other 2 treatments. Ingestion of conventional soy resulted in a flatus frequency (7.5 ± 1.9 times/12 h) that was significantly greater (P = 0.039) than that with rice (3.2 ± 0.8), whereas the frequency observed after ingestion of the low-oligosaccharide soy (3.9 ± 0.8) was comparable with that after rice (Figure 4). No abdominal symptoms differed significantly with either soy product from those observed with the rice control (Table 2). The ingestion of the meals did not result in a significant alteration in breath-methane concentrations from the zero time values, which averaged 18 ± 2.5 ppm. The sums of breath-methane concentrations for the 8 h after ingestion of the 3 meals were not significantly different in either study (data not shown).
View larger version (17K):
[in this window]
[in a new window]
|
FIGURE 4. Mean (± SEM) frequency of flatus passages in 20 healthy subjects in the 12 h after ingestion of puddings containing 80 g conventional soy concentrate or 80 g low-oligosaccharide soy concentrate or 80 g of a white rice cereal. Different letters indicate significant differences between groups, P < 0.05.
|
|
View this table:
[in this window]
[in a new window]
|
TABLE 2. Severity of symptoms over 12 h after ingestion of puddings containing 80 g of the soy concentrates or rice (control)1
|
|
|
DISCUSSION
|
|---|
The soluble oligosaccharides raffinose and stachyose are present in high concentrations in soybeans and other legumes (2, 11). These sugars consist of sucrose linked via -galactoside bonds to 1 or 2 galactose moieties to form raffinose and stachyose, respectively. The small intestine of humans (and animals) does not synthesize the -galactosidase required for the hydrolysis of these oligosaccharides to their constituent simple sugars. As a result, virtually 100% of ingested stachyose and raffinose reaches the colon. Colonic bacteria possess the enzyme systems necessary to ferment these sugars, and copious quantities of carbon dioxide and hydrogen may be released during the fermentation process. In addition to the well-known ability of soy and other legumes to cause gas in humans (1, 8, 12), soy products have been reported to induce digestive problems in animals (5). Soy is an inexpensive source of high-quality protein. In addition, recent studies have suggested that soy may have other beneficial features, including antineoplastic properties (13). Thus, a soy that lacks gas-producing factors could be useful for both humans and animals.
Recent studies have elucidated the metabolic pathway involved in raffinose and stachyose synthesis in soybeans (Figure 1
). At the end of this pathway, galactinol (1-O--D-galactopyranosyl-D-myo-inositol) serves as a galactose donor for reactions in which sucrose is converted to raffinose and raffinose to stachyose (14). These reactions are catalyzed by galactinol–sucrose galactosyltransferase and galactinol–raffinose galactosyltransferase, respectively. Beans with a naturally low activity of these enzymes have been identified and compared with conventional soybeans; these low-oligosaccharide soybeans are low in raffinose and stachyose but rich in galactinol (Table 1). The low concentration of the oligosaccharides presumably should reduce the gas-producing propensity of these beans. However, the fate of galactinol in the intestinal tract of humans and animals has not been studied. It seems likely that the small-intestinal mucosa will be impermeable to a water-soluble molecule the size of intact galactinol, which consists of galactose bound to myo-inositol. However, in the absence of information concerning the digestibility of galactinol and the ability of the colonic microflora to ferment nonabsorbed galactinol, the gas-producing propensity of a soybean low in oligosaccharides but rich in galactinol could not be predicted with certainty.
The purpose of the present study was to compare, in healthy human volunteers, gas production after ingestion of soy flour made from conventional soybeans with that after ingestion of a new variety of soybean that is naturally low in raffinose and stachyose. To this end, in double-blind, randomized, crossover studies, breath-hydrogen excretion and gaseous symptoms were assessed after ingestion of the 2 types of soy and rice.
Malabsorbed fermentable material is converted to hydrogen by multiple species of colonic bacteria. This hydrogen can be absorbed and excreted in the breath, passed through the rectum, or utilized by other bacterial species for the reduction of carbon dioxide to methane, or of sulfate to sulfide (15). Most of the hydrogen released during the slow fermentation of insoluble substrates such as fiber and resistant starch is utilized by other bacteria (16). However, an appreciable fraction of the hydrogen liberated during rapid fermentation of soluble carbohydrates—such as lactose, raffinose, and stachyose—is absorbed and excreted in the breath. Breath-hydrogen measurements have been shown to provide a semiquantitative assessment of the quantity of soluble carbohydrate reaching the colon (17).
Ingestion of conventional soy in quantities of 34 g (Figure 2) and 80 g (Figure 3) was associated with significantly greater increases in breath-hydrogen concentrations than those observed after ingestion of low-oligosaccharide soy. On the basis of these breath-hydrogen concentrations, it can be concluded that low-oligosaccharide soy contains less nonabsorbable, rapidly fermented substrate than does conventional soy. Thus, the advantage of low-oligosaccharide soy is that it produces less gas than does conventional soy; however, because it contains less nonabsorbable carbohydrate than conventional soy, the putative benefit to the colonic mucosa (18) of organic acids derived from nonabsorbable carbohydrate is lacking.
White rice has been shown to be the one complex carbohydrate that is nearly completely absorbed and hence results in minimal breath-hydrogen excretion (7). Although there appeared to be a trend toward greater hydrogen excretion after 80 g low-oligosaccharide soy than after 80 g white rice (Figure 3), this difference was not significant when assessed by ANOVA. However, this difference was significant (P = 0.026) when assessed by paired t test. Thus, it seems likely that some component of the low-oligosaccharide soy, such as galactinol or fiber, provided substrate that supported a small amount of hydrogen production in the colon.
The widespread belief that legume ingestion causes excessive rectal gas is supported by a large body of anecdotal evidence and many objective measurements (1, 8, 12). In the present investigation, no significant increase in flatus frequency was observed after ingestion of 34 g conventional soy, whereas ingestion of 80 g conventional soy (3.1 g raffinose and stachyose) resulted in a significant increase in flatus frequency. The relatively minor increase in flatus frequency induced by conventional soy flour (about 4 passages/12 h) was not totally unexpected given the relatively small quantity of oligosaccharides (3.06 g) in the 80-g dose. Christl et al (19) showed that as the rate of gas production in the colon increases, a smaller fraction of the gas is absorbed and a greater fraction is excreted through the rectum. Thus, it is possible that a slightly greater dose of conventional soy flour might have resulted in a disproportionate increase in rectal gas output and a more readily recognizable increase in flatus frequency.
Techniques to reduce the raffinose and stachyose contents of soybean products have been the subject of extensive investigation. Cooking does not eliminate these heat-stable oligosaccharides; however, germination for 1–4 d results in the utilization of the bulk of these sugars (20). Incubation with various microorganisms or enzymes derived from microorganisms yields products with a markedly reduced oligosaccharide content (21, 22). Ultrafiltration of the aqueous fraction of soybeans has also been successful in removing oligosaccharides from soybeans (23). The present study showed that soy seeds naturally low in oligosaccharides produced little flatulence, significantly less than that associated with conventional soy. Thus, this new seed may represent a useful alternative to conventional soybeans.
|
FOOTNOTES
|
|---|
1 From the Minneapolis Veterans' Affairs Medical Center.
2 Supported in part by the Department of Veterans' Affairs; the National Institute of Diabetes and Digestive and Kidney Diseases (RO1-DK-13093); Optimum Quality Grains, Des Moines, IA; and DuPont Protein Technologies International, St Louis.
3 Address reprint requests to MD Levitt, Research Office (151), Minneapolis Veterans' Affairs Medical Center, One Veterans Drive, Minneapolis, MN 55417. E-mail: levitt.michael(stf){at}minneapolis.va.gov.
|
REFERENCES
|
|---|
-
Steggerda FR, Richards EA, Rackis JJ. Effects of various soybean products on flatulence in the adult man. Proc Soc Exp Biol Med 1966;121:1235–9.[Medline]
-
Rackis JJ, Honig DH, Sessa DJ, Steggerda FR. Flavor and flatulence factors in soybean protein products. J Agric Food Chem 1970;18:977–82.[Medline]
-
Savitri A, Desikachar HSR. A comparative study of flatus production in relation to the oligosaccharide content of some legumes. Nutr Rep Int 1985;31:337–44.
-
Olson AC, Gray GM, Gumbmann MR, Sell CR, Wagner JR. Flatus causing factors in legumes. In: Ory RL, ed. Antinutrients and natural toxicants in foods. Westport, CT: Food and Nutrition Press, 1981:275–94.
-
Coon CN, Leske KL, Akavanichan O, Cheng TK. Effect of oligosaccharide-free soybean meal on true metabolizable energy and fiber digestion in adult roosters. Poult Sci 1990;69:787–93.[Medline]
-
Suarez FL, Savaiano DA, Levitt MD. A comparison of symptoms in people with self-reported severe lactose intolerance after drinking milk or lactose-hydrolyzed milk. N Engl J Med 1995;333:1–4.[Abstract/Free Full Text]
-
Levitt MD, Hirsh P, Fetzer CA, Sheahan M, Levine AS. H2 excretion after ingestion of complex carbohydrates. Gastroenterology 1987;92:383–9.[Medline]
-
Suarez FL, Furne JK, Springfield JR, Levitt MD. Insights into human colonic physiology obtained from study of flatus composition. Am J Physiol 1997;272:G1028–33.[Abstract/Free Full Text]
-
SYSTAT Inc. SYSTAT for the Macintosh, version 5.2. Evanston, IL: SYSTAT Inc, 1992.
-
SPSS Inc. SPSS advance statistics, version 6.1. Chicago: SPSS Inc, 1994.
-
Rackis JJ. Flatulence caused by soy and its control through processing. J Am Oil Chem Soc 1981;58:503–9.
-
Steggerda FR, Dimmick JF. Effect of bean diet on concentration of carbon dioxide in flatus. Am J Clin Nutr 1966;19:120–4.[Abstract]
-
Rao AV, Sung MK. Saponins as anticarcinogens. J Nutr 1995; 125:717S–24S.
-
Kabat EA, MacDonald DL, Ballou CE, Fisher HOL. On the structure of galactitol. J Am Chem Soc 1953;75:4507–9.
-
Gibson GR, Cummings JH, MacFarlane GT, et al. Alternative pathways for hydrogen disposal during fermentation in the human colon. Gut 1990;31:679–83.[Abstract/Free Full Text]
-
Strocchi A, Ellis CJ, Levitt MD. Use of metabolic inhibitors to study H2 consumption by human feces: evidence for undescribed metabolic pathways. J Lab Clin Med 1992;120:320–7.
-
Bond JH, Levitt MD. Use of pulmonary hydrogen (H2) measurements to quantitate carbohydrate malabsorption: study of partially gastrectomized patients. J Clin Invest 1972;51:1219–25.
-
Hassig CA, Tong JK, Schreiber SL. Fiber-derived butyrate and the prevention of colon cancer. Chem Biol 1997;4:783–9.[Medline]
-
Christl SU, Murgatroyd PR, Gibson GR, Cummings JH. Production, metabolism, and excretion of hydrogen in the large intestine. Gastroenterology 1992;102:1269–77.[Medline]
-
East JW, Nakayama TOM, Parkman SB. Changes in stachyose, sucrose, and monosaccharides during germination of soybeans. Crop Sci 1972;12:7–9.
-
Mittal BK, Steinkraus KH. Utilization of oligosaccharides by lactic acid bacteria during fermentation of a soy milk. J Food Sci 1975;40:114–8.
-
Cruz R, Batistela JC, Wosiaski G. Microbial alpha-galactosidase for soy milk processing. J Food Sci 1982;46:1196–200.
-
Omosaiye O, Cheryan M, Mathews ME. Removal of oligosaccharides from soybean water extracts by ultrafiltration. J Food Sci 1978;43:354–60.
Received for publication March 13, 1998.
Accepted for publication June 4, 1998.
This article has been cited by other articles:
|
|
|
|
 
J. D. Neus, W. R. Fehr, and S. R. Schnebly
Agronomic and Seed Characteristics of Soybean with Reduced Raffinose and Stachyose
Crop Sci.,
February 23, 2005;
45(2):
589 - 592.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
Patrick De Boever, B. Deplancke, and W. Verstraete
Fermentation by Gut Microbiota Cultured in a Simulator of the Human Intestinal Microbial Ecosystem Is Improved by Supplementing a Soygerm Powder
J. Nutr.,
October 1, 2000;
130(10):
2599 - 2606.
[Abstract]
[Full Text]
|
|
|
TOP
ABSTRACT
INTRODUCTION
), 34 g low-oligosaccharide soy concentrate (O), and lactose-hydrolyzed cow milk (control, 
). The sum of the hourly breath-hydrogen concentrations from 0 to 8 h was significantly greater (P = 0.005) after ingestion of conventional soy than after ingestion of low-oligosaccharide soy or lactose-hydrolyzed milk.