in expts 1 and 2 , dz ( purity > 98  % ; nagara science co. ltd ) was added at 1  g / kg 
 instead of sucrose to the control diet ( table 
 1 ) . catechin ( egcg  >  90  % , dsm nutrition k. k. ) at 1  g / kg was added instead 
 of maize starch to the control diet in expt 1 . pd ( wako pure chemical industries ltd ) or 
 raf ( d ( + ) -raf pentahydrate ; wako pure chemical industries ltd ) at 50  g / kg was added 
 instead of maize starch to the control diet in expt 2 . table 1.composition of the experimental diet ( g / kg diet)*ingredientcontroldzdz  +  catecatedz  +  pdpddz  +  rafrafmaize starch52955295519551954795479547954795casein200200200200200200200200sucrose10099991009910099100maize oil7070707070707070cellulose5050505050505050mineral mixture*3535353535353535vitamin mixture*1010101010101010l - cystine33333333choline bitartrate2525252525252525tert - butylhydroquinone00140014001400140014001400140014daidzein10101010catechin1010polydextrose5050raffinose5050control , control diet ; dz , daidzein supplemented diet ; dz  +  cate , dz and 
 catechin - supplemented diet ; cate , cate - supplemented diet ; dz  +  pd , dz and 
 polydextrose - supplemented diet ; pd , pd - supplemented diet ; dz  +  raf , dz and 
 raffinose - supplemented diet ; raf , raf - supplemented diet.*prepared according to the ain-93  g formulation.experiment 1.experiment 2 . composition of the experimental diet ( g / kg diet ) * control , control diet ; dz , daidzein supplemented diet ; dz  +  cate , dz and 
 catechin - supplemented diet ; cate , cate - supplemented diet ; dz  +  pd , dz and 
 polydextrose - supplemented diet ; pd , pd - supplemented diet ; dz  +  raf , dz and 
 raffinose - supplemented diet ; raf , raf - supplemented diet . prepared according to the ain-93  g formulation . female mice of the ddy strain , aged 8 weeks , were purchased from the shizuoka 
 laboratory animal center . they were housed in individual cages in a temperature- and 
 humidity - controlled room ( 23    1c and 60    5  % relative humidity ) with a 12  h 
 light12  h dark cycle . they were given free access to an american institute of nutrition 
 ( ain)-93 g diet with maize oil instead of soyabean oil for 4  d before 
 surgery 
 . mice were either sham - operated ( sham ) or ovx on the same day . the 
 ovx mice were randomly divided into four groups ( n 68 ) : ovx - control 
 ( ovx ) , ovx fed a 01  % dz - supplemented diet ( ovx  +  dz ) , ovx fed a 01  % dz- and 1  % 
 catechin - supplemented diet ( ovx  +  dz  +  cate ) and ovx fed 1  % catechin - supplemented diet 
 ( ovx  +  cate ) . table 1 shows the composition of 
 the experimental diets , which were prepared according to the ain-93  g 
 formulation 
 . maize oil was used to eliminate any possible contamination from the 
 isoflavones in soyabean oil . mice were pair - fed their respective diets for 42  d with 
 free access to distilled water during this period . after 42  d of treatment , the mice 
 were fasted overnight and then euthanised by exsanguination under anaesthesia , weighed 
 and blood was drawn and stored at 80c until assay . the contents were collected , the caecal tissue 
 was washed with saline and the weight of the contents was calculated by subtracting the 
 tissue weight from the total weight . the ph of the caecal contents was measured with a 
 ph meter ( model b-212 , twin compact ph meter , horiba , scientific ltd ) . all experimental procedures involving 
 animals were approved by the national institute of health and nutrition guidelines for 
 the care and use of laboratory animals . mice were acclimatised and either sham - operated or ovx on the same day , as in expt 1 . the mice were divided into seven groups ( n 68 ) : sham - operated ( sham ) 
 or ovx - control ( ovx ) , ovx fed a 01  % dz - supplemented diet ( ovx  +  dz ) , ovx fed a 01  % 
 dz5  % pd - supplemented diet ( ovx  +  dz  +  pd ) , ovx fed a 5  % pd - supplemented diet 
 ( ovx  +  pd ) , ovx fed a 01  % dz5  % raf - supplemented diet ( ovx  +  dz  +  raf ) and ovx fed a 
 5  % raf - supplemented diet ( ovx  +  raf ) . the mice were pair - fed their respective diets 
 ( table 1 ) for 42  d with free access to 
 distilled water during this period . after 41  d of treatment , 24-h urine was collected 
 and stored at 20c until assay . at the end of the experiment , body weight , uterus 
 weights , caecal weights and caecal ph were measured and the caecal contents , right femur 
 and blood were collected as described in expt 1 . femoral bmd was measured by dual - energy x - ray absorptiometry ( dxa ; model dcs-600ex - r , 
 aloka ltd ) . the scanned area 
 of the mouse femur was divided into three equal parts of 53  mm each : the proximal 
 femur , midshaft and distal femur . plasma and urinary dz and equol were analysed by the time - resolved fluoroimmunoassay 
 methods of wang et al . and urine and plasma were hydrolysed by glucuronidase and sulphatase 
 after the plasma was extracted using ether . plasma and urinary dz and equol 
 concentrations were determined by fluorescence using a delfia victor 1420 multilabel 
 counter ( perkin elmer co. ltd ) . the final results  =  concentration 
 ( read )    1/recovery    dilution factor ( nmol / l ) . activity of -glucosidase was indicated as the rate of release of 
 p - nitrophenol from p - nitrophenylglucoside . the 
 reaction mixture contained 01  ml of a 5  mmol / l substrate solution and 02  ml of a 1:20 
 ( v / v ) dilution of the caecal sample in 01  mm - phosphate buffer at ph 64 . the 
 reaction mixture was incubated for 30  min at 37c , and the 
 p - nitrophenol concentration was measured spectrophotometrically at 
 400  nm after the addition of 16  ml of 025  m - sodium carbonate . enzyme activity 
 was expressed as mmol product hydrolysed per whole caecal contents in 30  min . the data are expressed as mean values with their standard errors of the difference 
 between means . the significance of differences in bmd was determined by one - factor anova 
 and fisher 's protected least significant difference test ( spss version 11.0 ; spss inc . ) . body weight was used as a covariate in the analysis of bmd to adjust for possible 
 confounding effects . initial body weights of all groups of mice did not differ significantly from each other 
 ( data not shown ) . final body weights were significantly lower in the ovx  +  dz  +  cate and 
 ovx  +  cate groups ( 299 ( se 07 ) and 305 ( se 07 )  g , respectively ) than 
 in the sham , ovx and ovx  +  dz groups ( 351 ( se 15 ) , 370 ( se 09 ) and 
 347 ( se 07 )  g , respectively ) . the uterine weight was lower in ovx mice than in 
 sham - operated mice , and treatment with dz or catechin did not affect the uterine weight in 
 ovx mice . plasma dz concentrations were significantly increased by dz feeding in the 
 ovx  +  dz and the ovx  +  dz  +  cate groups ( 0927 ( se 0031 ) , 0863 ( se 
 0025 )  mol / l , respectively ) . plasma equol was detected in the ovx  +  dz group ( 967 
 ( se 117 ) mol / l ) , but it was not detected in the ovx  +  dz  +  cate group . the wet weight of caecal contents was significantly higher in the ovx  +  cate group ( 0867 
 ( se 0118 )  g/100  g body weight ) than in the ovx group ( 0448 ( se 
 0074 )  g/100  g body weight ) , and tended to be higher in the ovx  +  dz  +  cate group ( 0608 
 ( se 0092 )  g/100  g body weight ) than in the ovx group 
 ( p  =  0276 ) . there were no significant differences in caecal ph and 
 -glucosidase activity between all groups ( data not shown ) . femoral bmd was significantly lower in ovx mice than in sham mice ( table 2 ) , and dz treatment inhibited femoral bone loss . however , the 
 femoral bmd in the ovx  +  dz  +  cate group was lower than that in the ovx  +  dz group . the 
 inhibition of bone loss by dz was attenuated by treatment with catechin in ovx mice . table 2.bone mineral density ( bmd ) of the femur obtained from the mice in expt 1(mean values with their standard errors)shamovxovx +  dzovx  +  dz  +  cateovx  +  catemean 
 se 
 mean 
 se 
 mean 
 se 
 mean 
 se 
 mean 
 se 
 whole femur bmd ( mg / cm)4011133907379083280335109proximal region ( mg / cm)4410740207440113810839711middle region ( mg / cm)3510831207346103070333210distal region ( mg / cm)4741934903405133550336811sham , sham - operated mice fed the control diet ; ovx , ovariectomised mice fed the 
 control diet ; ovx  +  dz , ovx mice fed the 01  % daidzein - supplemented diet ; 
 ovx  +  dz  +  cate , ovx mice fed a combination of 01  % dz and 1  % 
 catechin - supplemented diet ; ovx  +  cate , ovx mice fed the 1  % cate - supplemented 
 diet . mean values within a row with unlike superscript letters were 
 significantly different ( p < 005 ) . bone mineral density ( bmd ) of the femur obtained from the mice in expt 1 ( mean values with their standard errors ) sham , sham - operated mice fed the control diet ; ovx , ovariectomised mice fed the 
 control diet ; ovx  +  dz , ovx mice fed the 01  % daidzein - supplemented diet ; 
 ovx  +  dz  +  cate , ovx mice fed a combination of 01  % dz and 1  % 
 catechin - supplemented diet ; ovx  +  cate , ovx mice fed the 1  % cate - supplemented 
 diet .   mean values within a row with unlike superscript letters were 
 significantly different ( p < 005 ) . the initial and final body weights of all groups of mice did not differ significantly 
 from each other ( data not shown ) . treatment with dz , pd and/or raf did not affect the 
 uterine weight in ovx mice . the plasma dz concentration was significantly higher in the 
 ovx  +  dz group than in the other groups ( fig . the plasma equol concentrations were higher in the ovx  +  dz  +  raf and 
 ovx  +  dz  +  pd groups ( 467 ( se 089 ) and 493 ( se 049 )  mol / l , 
 respectively ) than in the ovx  +  dz group ( 028 ( se 054 ) 1(b ) ) . the urinary dz and equol concentrations in 
 dz - fed groups were significantly higher than in the other groups ( fig . 2(a ) and ( b ) ) . the ratio 
 of urinary equol to dz concentration tended to be higher in the ovx  +  dz  +  raf group than 
 in the ovx  +  dz group ( p  =  0124 ) , and the ratio was significantly higher 
 in the ovx  +  dz  +  pd group than in the ovx  +  dz group ( p  =  007 ) . 1.plasma concentrations of daidzein ( dz ) ( a ) and equol ( b ) in mice in expt 2 . sham , 
 sham - operated mice fed the control diet ; ovx , ovariectomised mice fed the control 
 diet ; ovx  +  dz , ovx mice fed the 01 % dz - supplemented diet ; ovx  +  dz  +  pd , ovx mice 
 fed a combination of 01 % dz and 5 % polydextrose - supplemented diet ; ovx  +  pd , ovx 
 mice fed the 5 % polydextrose - supplemented diet ; ovx  +  dz  +  raf , ovx mice fed a 
 combination of 01 % dz and 5 % raffinose - supplemented diet ; ovx  +  raf , ovx mice fed 
 the 5 % raf - supplemented diet . .   mean values with unlike letters were 
 significantly different ( p  <  005 ) . 2.concentrations of urinary daidzein ( dz ) ( a ) and equol ( b ) and the ratio of urinary 
 equol : daidzein concentrations ( c ) in mice in expt 2 . sham , sham - operated mice fed 
 the control diet ; ovx , ovariectomised mice fed the control diet ; ovx  +  dz , ovx mice 
 fed the 01 % daidzein - supplemented diet ; ovx  +  dz  +  pd , ovx mice fed a combination 
 of 01 % dz and 5 % polydextrose - supplemented diet ; ovx  +  pd , ovx mice fed 5 % 
 polydextrose - supplemented diet ; ovx  +  dz  +  raf , ovx mice fed a combination of 01 % 
 dz and 5 % raffinose - supplemented diet ; ovx  +  raf , ovx mice fed 5 % raf - supplemented 
 diet . . 
  mean values with unlike letters were significantly different 
 ( p  <  005 ) . plasma concentrations of daidzein ( dz ) ( a ) and equol ( b ) in mice in expt 2 . sham , 
 sham - operated mice fed the control diet ; ovx , ovariectomised mice fed the control 
 diet ; ovx  +  dz , ovx mice fed the 01 % dz - supplemented diet ; ovx  +  dz  +  pd , ovx mice 
 fed a combination of 01 % dz and 5 % polydextrose - supplemented diet ; ovx  +  pd , ovx 
 mice fed the 5 % polydextrose - supplemented diet ; ovx  +  dz  +  raf , ovx mice fed a 
 combination of 01 % dz and 5 % raffinose - supplemented diet ; ovx  +  raf , ovx mice fed 
 the 5 % raf - supplemented diet . mean values with unlike letters were 
 significantly different ( p  <  005 ) . concentrations of urinary daidzein ( dz ) ( a ) and equol ( b ) and the ratio of urinary 
 equol : daidzein concentrations ( c ) in mice in expt 2 . sham , sham - operated mice fed 
 the control diet ; ovx , ovariectomised mice fed the control diet ; ovx  +  dz , ovx mice 
 fed the 01 % daidzein - supplemented diet ; ovx  +  dz  +  pd , ovx mice fed a combination 
 of 01 % dz and 5 % polydextrose - supplemented diet ; ovx  +  pd , ovx mice fed 5 % 
 polydextrose - supplemented diet ; ovx  +  dz  +  raf , ovx mice fed a combination of 01 % 
 dz and 5 % raffinose - supplemented diet ; ovx  +  raf , ovx mice fed 5 % raf - supplemented 
 diet . . 
  mean values with unlike letters were significantly different 
 ( p  <  005 ) . wet weights of the caecal contents in the ovx  +  dz  +  pd and ovx  +  dz  +  raf groups were 
 higher than those in the sham , ovx , ovx  +  dz and ovx  +  pd groups ( table 3 ) . caecal ph was lower in the ovx  +  dz  +  pd group than in the 
 other groups ( table 3 ) . caecal -glucosidase 
 activity was significantly higher in the ovx  +  dz  +  pd group than in the other groups 
 ( table 3 ) . the wet weights of the caecal 
 contents were elevated in mice fed dz and raf , but caecal ph and caecal -glucosidase 
 activity were not affected by these additives . table 3.wet weights of the caecal content , ph and -glucosidase activity in the mice in 
 expt 2(mean values with their standard errors)shamovxovx +  dzovx  +  dz  +  pdovx  +  pdovx  +  dz  +  rafovx  +  rafmean 
 se 
 mean 
 se 
 mean 
 se 
 mean 
 se 
 mean 
 se 
 mean 
 se 
 mean 
 se 
 wet weight of the caecal contents*01540015012300200180001702920013016400240248002902020028caecal ph787007768011777008691016740005757004770007caecal -glucosidase activity01990035036900960256006547631830101603620493016202010021sham , sham - operated mice fed the control diet ; ovx , ovariectomised mice fed the 
 control diet ; ovx  +  dz , ovx mice fed 01  % daidzein - supplemented diet ; 
 ovx  +  dz  +  pd , ovx mice fed a combination of 01  % dz and 5  % 
 polydextrose - supplemented diet ; ovx  +  pd , ovx mice fed 5  % pd - supplemented diet ; 
 ovx  +  dz  +  raf , ovx mice fed a combination of 01  % dz and 5  % 
 raffinose - supplemented diet ; ovx  +  raf , ovx mice fed 5  % raf - supplemented 
 diet . mean values within a row with unlike superscript letters were 
 significantly different ( p < 005).*g/100  g body weight.hydrolysed product ( mmol)/whole caecal contents per 30  min . wet weights of the caecal content , ph and -glucosidase activity in the mice in 
 expt 2 ( mean values with their standard errors ) sham , sham - operated mice fed the control diet ; ovx , ovariectomised mice fed the 
 control diet ; ovx  +  dz , ovx mice fed 01  % daidzein - supplemented diet ; 
 ovx  +  dz  +  pd , ovx mice fed a combination of 01  % dz and 5  % 
 polydextrose - supplemented diet ; ovx  +  pd , ovx mice fed 5  % pd - supplemented diet ; 
 ovx  +  dz  +  raf , ovx mice fed a combination of 01  % dz and 5  % 
 raffinose - supplemented diet ; ovx  +  raf , ovx mice fed 5  % raf - supplemented 
 diet . mean values within a row with unlike superscript letters were 
 significantly different ( p < 005 ) . femoral bmd of the whole and each of the three parts was significantly lower in the ovx 
 group than in the sham group ( p  <  005 ) ( fig . the dz treatment groups showed inhibited femoral bone 
 loss compared with the ovx , ovx  +  pd and ovx  +  raf groups . the bmd of the whole , proximal , middle and distal regions of 
 the femur in the ovx  +  dz  +  pd and ovx  +  dz  +  raf groups was significantly higher than 
 that in the ovx group ( fig . 3(a)(d ) ) . 
 furthermore , the bmd of the whole , middle and distal femur was significantly higher in the 
 ovx  +  dz  +  pd group than in the ovx  +  dz group ( fig . 3(a ) , ( c ) and ( d ) ) , and tended to be higher in the ovx  +  dz  +  raf group than in the 
 ovx  +  dz group ( fig . the bmd 
 of the distal femur was significantly higher in the ovx  +  dz  +  pd group than in the 
 ovx  +  dz  +  raf group ( p  =  0038 ) ( fig . whole femur 
 bmd ( a ) , proximal femur bmd ( b ) , middle femur bmd ( c ) and distal femur bmd ( d ) . sham , sham - operated mice fed the control diet ; ovx , ovariectomised mice fed the 
 control diet ; ovx  +  dz , ovx mice fed 01 % daidzein - supplemented diet ; 
 ovx  +  dz  +  pd , ovx mice fed a combination of 01 % dz and 5 % 
 polydextrose - supplemented diet ; ovx  +  pd , ovx mice fed 5 % polydextrose - supplemented 
 diet ; ovx  +  dz  +  raf , ovx mice fed a combination of 01 % dz and 5 % 
 raffinose - supplemented diet ; ovx  +  raf , ovx mice fed 5 % raf - supplemented diet . . 
  mean values with unlike letters were significantly different 
 ( p  <  005 ) . whole femur 
 bmd ( a ) , proximal femur bmd ( b ) , middle femur bmd ( c ) and distal femur bmd ( d ) . sham , sham - operated mice fed the control diet ; ovx , ovariectomised mice fed the 
 control diet ; ovx  +  dz , ovx mice fed 01 % daidzein - supplemented diet ; 
 ovx  +  dz  +  pd , ovx mice fed a combination of 01 % dz and 5 % 
 polydextrose - supplemented diet ; ovx  +  pd , ovx mice fed 5 % polydextrose - supplemented 
 diet ; ovx  +  dz  +  raf , ovx mice fed a combination of 01 % dz and 5 % 
 raffinose - supplemented diet ; ovx  +  raf , ovx mice fed 5 % raf - supplemented diet . . 
  mean values with unlike letters were significantly different 
 ( p  <  005 ) . in the present study , we examined the effects of two of the ingredients that form part of 
 the japanese diet , catechin and dietary fibre , on equol production and bmd in dz - treated ovx 
 mice . dietary fibre stimulated equol production and enhanced the bone - protective effects of 
 dz on femoral bone . on the other hand , catechin reduced serum equol levels and suppressed 
 the beneficial effect of dz on femoral bone loss . the data suggest that the bioavailability 
 of isoflavones may be altered by dietary fibre , in particular , pd . in our study , the wet weight of caecal contents in the catechin feeding group was higher 
 than that in the ovx group , but there were no significant differences in caecal ph and 
 -glucosidase activity between all groups ( data not shown ) . furthermore , a significant 
 decrease in the concentration of serum equol was observed in these groups . it has been 
 reported that the polyphenol egcg in green tea has therapeutic effects on colon 
 diseases 

 . shirakami et al . reported that drinking water 
 containing egcg was beneficial to the suppression of cancer development in the inflamed 
 colon in mice 
 . additionally , the ameliorative effect of egcg on inflammatory bowel 
 disease has been observed in rats 
 . these studies adopted drinking water or diet containing 001020  % 
 egcg . in our study , supplementation of catechin at 1  % of the diet resulted in negative 
 effects reducing the equol production in the intestine . catechin is one of the polyphenol 
 compounds ; polyphenols are potential modulators of digestive fermentation 

 . molan et al . reported that orally administering rats 
 with green tea extract for 6  d resulted in a slight increase in the numbers of caecal 
 lactobacilli and bifidobacteria but decreased the numbers of bacteroides 
 and clostridia significantly 
 . reported that 
 supplementation with 03  % polyphenol extracts significantly increased the caecal content 
 weight 
 , but did not measure caecal bacteria . available data have suggested 
 that these compounds are a diarrhoeal 

  or an anti - diarrhoeal agent 
 . in our study , moisture content of caecal samples was not measured ; 
 further investigations including those of water absorption in the intestine are needed . furthermore , in our study , the dietary 1  % catechin level was higher than in previous 
 reports 

 . hara et al . also reported that a diet supplemented 
 with 02  % tea polyphenols significantly decreases the total bacteria level in 
 pigs 
 . equol is metabolised by intestinal bacteria ; thus , a decrease in the 
 abundance of bacteria in the intestine may lead to a decrease in equol production . these 
 results suggested that a large amount of catechin may alter the intestinal environment and 
 depress equol production in the intestine . the catechin in green tea , ( )-egcg , increases the osteogenic function of mesenchymal 
 stem cells 
 , and animal studies have strongly suggested that green tea prevents 
 bone loss 

 . in contrast with previous reports , we found that the inhibition of 
 bone loss by dz was attenuated by catechin treatment ( table 2 ) . the discrepancy between previous reports and the present study may be 
 due to a different dosage of catechin . other experiments examining the effects of catechin 
 on animal models adopted drinking water containing 01 or 05  % green tea 
 catechin 

 . shen et al . reported that 05  % catechin 
 supplemented in drinking water of rats attenuated trabecular and cortical bone loss by 
 increasing bone formation 
 . in contrast , iwaniec et al . reported that bone 
 mineral content and volumetric bmd were reduced by green tea extract consumption in 
 growing male mice 
 . in that study , the diets contained green tea extract at 1 or 2  % of 
 the diet . thus , a high dose of catechin may reduce bone mass and/or accelerate bone loss . further studies are needed to confirm the effects of catechin extracts on bone 
 metabolism . our results showed that pd had a greater effect on equol production and on the enteric 
 environment such as ph and weight of the caecal contents ( table 3 ) . the urinary equol concentration and its ratio to dz 
 concentration were significantly higher in the ovx  +  dz  +  pd group than in the ovx  +  dz 
 group in expt 2 ( fig . acidification of caecal 
 content was more pronounced with the dz- and pd - containing diet , compared with pd alone 
 ( table 3 ) . the data were consistent with a 
 previous report that feeding of rats with 03  % flavonoid and 5  % inulin , a kind of 
 dietary fibre added to the diet , induced significant enlargement of the caecum compared 
 with rats fed the control diet . the greatest enlargement of caecal contents was observed 
 in rats fed a flavonoid - containing diet 

 . since dz is a kind of flavonoid , it is possible that it induces 
 synergistic effects with pd on the enteric environment . these results suggest that the 
 combination of dietary fibres , either pd or raf , and dz enhances not only the metabolism 
 of isoflavone conjugates to aglycone but also the metabolism of aglycone to equol from dz . further studies are required to confirm the effects of each type of dietary fibre on equol 
 production . the ability to produce equol depends on the presence of certain intestinal microflora ; 
 thus , the enteric environment has been known to affect equol production 
 . we measured -glucosidase activity in the caecal contents and found 
 an increase in enzymatic activity in mice fed pd and dz ( table 3 ) . the activity of -glucosidase in the caecal contents is an 
 indicator of the activity of the intestinal enzymes hydrolysing the glycoside bond of 
 isoflavone conjugates and this leads to stimulation of the intestinal absorption of 
 isoflavone aglycones . ohta et al . reported that fructo - oligosaccharide increases 
 -glucosidase activity in mice 
 . pd produces scfa in the process of fermentation and has a stimulatory 
 effect on colonic lactobacillus and bifidobacterium 
 species similar to fructo - oligosaccharides 

 . jie et al . reported that ingestion of pd increases 
 the faecal weight and faecal abundance of lactobacillus and 
 bifidobacterium species but decreases faecal ph in chinese 
 subjects 
 . similarly , raf produces scfa and increases the faecal populations of 
 bifidobacterium and lactobacillus 
 species 
 . lactobacillus and bifidobacterium species have been 
 identified to play a role in the metabolism of dz to equol 

 . we have previously reported that a diet supplemented with dz and 
 resistant starch , which is not absorbed in the intestine , increases the equol production 
 to occupation ratio of bifidobacterium species to that of the intestinal 
 microflora in mice 
 . although we did not measure the composition of the intestinal 
 microflora , it is possible that lactobacillus and/or 
 bifidobacterium species increased in the present study . we were able to show that ovx mice whose diets were supplemented with dz or a combination 
 of dz and pd or raf maintained their femoral bmd compared with ovx mice ( fig . in particular , mice fed dz  +  pd maintained 
 their bmd more effectively than mice fed dz  +  raf . although pd and raf improve caecal ca 
 absorption in rats 

 , a diet supplemented with pd or raf alone did not maintain the bmd in 
 ovx mice in our study . these results are similar to those of our previous study in which 
 diets supplemented with resistant starch and dz efficiently prevented bmd decrease in ovx 
 mice 
 . in the present study , the synergistic effects may be credited with 
 the ability of dietary fibre to enhance equol production . the plasma equol 
 concentrations in the ovx  +  dz group in expt 1 and those in expt 2 were 967 ( se 
 117 ) and 285 ( se 151 )  mol / l , respectively . the plasma equol concentration in 
 the ovx  +  dz group in expt 1 was approximately 3-fold higher than that in the ovx  +  dz 
 group in expt 2 . as the reason for this difference , serum equol concentration is 
 influenced by the timing of blood collection and changes substantially 
 . therefore , equol may have reached peak concentrations in plasma and 
 may have been excreted in the urine . poulsen et al . reported that 491  % 
 of dz and its metabolites , including equol , were excreted in the urine and faeces of rats 
 within 24  h 
 . precursor relationship as the concentration ratio of equol to dz in urine is a 
 more reliable indicator of the amount of dz converted to equol 
  ( fig . this study demonstrated the effects of components from a typical japanese diet 
 ( isoflavones , tea catechin or dietary fibre ) on equol production and bone metabolism in ovx 
 mice . this 
 effect was greater than that of dz alone for preventing bone loss in dz - treated ovx mice . in 
 contrast , catechin reduced serum equol levels and suppressed the beneficial effect of dz on 
 femoral bone loss . these results suggest that additional studies should be carried out to 
 determine whether a combination of isoflavones and dietary fibre is useful in maintaining 
 the bone health of postmenopausal women .
equol is a metabolite of the isoflavone daidzein ( dz ) and is produced by the bacterial 
 microflora in the distal intestine and colon . 
 some epidemiological studies have reported 
 an association between increased equol production and intakes of green tea or dietary 
 fibre , which are ingredients of the standard japanese diet . 
 we examined the effects of a 
 diet supplemented with dz and tea catechin or dietary fibre on equol production and bone 
 mineral density in ovariectomised ( ovx ) mice . 
 female mice of the ddy strain were either 
 sham operated or ovx . 
 ovx mice were fed a control diet , a 01  % dz - supplemented diet or a 
 01  % dz diet supplemented with one of the food components commonly consumed in the 
 japanese diet . 
 the mice were given 1  % tea catechin ( w / w ) as part of the diet in expt 1 or 
 5  % polydextrose ( pd ) and 5  % raffinose ( raf ) ( w / w ) as part of the diet in expt 2 . 
 catechin reduced serum equol levels and attenuated the beneficial effect of dz on femoral 
 bone loss . 
 the soluble dietary fibres pd and raf stimulated equol production , and enhanced 
 the bone - protective effects of dz on femoral bone . 
 these results suggest that dietary 
 fibre , in particular , pd , may alter the bioavailability of isoflavones and prevent 
 osteopenia in ovx mice .