30-1
Bioavailability of Ca Fortifi ed 7S Globulin Rich Soy Milk in Young Women
Chie TARUMIZU, Noriko WAKIKAWA, Hien Vu THITHU, Nobuko SARUKURA,
Andrea WAKITA and Shigeru YAMAMOTO
Ochanomizu University, Graduate School of Humanities & Sciences, Tokyo 112-8610
ABSTRACT
There are contradictions in the understanding of the bioavailability of food Ca. The
Ca concentration of TOFU is as high as that of cow's milk, while bioavailability of
the former is thought to be lower than that of the latter according to a report from
an older study in Japan; however the studies from all over the world have shown
rather similar absorption rates for TOFU, soybean products, cow's milk and dairy
product. In addition, there are remarkable differences in the recommendation of
Ca intake world-wide, which also suggests inconsistency in Ca bioavailability. The
purpose of this study was to establish the absorption rates and requirements of Ca
in Ca-fortifi ed soy milk and cow's milk by the improved balance studies. Twelve
young female subjects were randomly assigned to two groups and were given
basal diets (Ca 300 mg/d) with Ca fortifi ed soy milk (Ca 300 mg/d) or cow's milk
(Ca 300 mg/d) for 15 d separated by a 2-6 wk washout interval in a cross-over
design. Basal diets were 5 d cycle menu. Feces, urine and diets of the last 5 d were
collected in both dietary periods and were digested by the microwave method and
Ca concentration was analyzed by the atomic absorption method. The apparent
absorption rates were 19% in both the soy milk and cow's milk groups. The present
study showed that the Ca absorption rates for soy milk and cow's milk were similar
in young female subjects.
Soy Protein Research, Japan 12, 1-10, 2009.
Key words : calcium, soy milk, cow's milk, bioavailability, women
30-2
Systematic Review and Verifi cation of Preventive Effects of
Soy Constituents on Lifestyle-Related Diseases:
Isolated Soy Isoflavone Supplements for Postmenopausal Bone Loss:
Meta-Analysis of Randomized Controlled Trials (Part II)
Yoshiko ISHIMI1, Kyoko TAKU2, Keizo UMEGAKI3 and Jun YAMAUCHI1
1Project for Bio-index, Nutritional Epidemiology Program,
2Project for Health Communication,
3Project for information Network of Health Food, Information Center,
National Institute of Health and Nutrition, Tokyo 162-8636
ABSTRACT
This study was intended to 1) examine the Effects of the functional components of
soy foods on prevention of life style-related diseases via a systematic review and
meta-analysis of the related articles, 2) survey the labeling and the contents of
functional components of soy in the health foods. This year, we focused on soybean
Isoflavones and soy protein. This meta-analysis was performed to clarify the eff ect
of isolated soy Isoflavone supplements alone on postmenopausal bone loss. PubMed,
CENTRAL, ICHUSHI, CNKI, Wanfang Data, CQVIP, and NSTL up to September
2008 were searched for articles of randomized placebo-controlled trials published in
English, Japanese, or Chinese reporting the outcomes of Isoflavone supplements on
lumbar spine or hip (including femoral neck, hip total, and trochanter) bone mineral
density (BMD) in postmenopausal women. Meta-analysis of six high-quality trials
including 712 women with low bone mass, revealed significant overall weighted
mean differences of 22.05 mg/cm2 (95% CI: 1.62 to 42.48; p =0.03), or 2.49% (95% CI:
0.23 to 4.76; p =0.03) in change of spine BMD from baseline between participants
ingesting an average of 65 mg soy Isoflavones/d (in aglycone form) and participants
ingesting placebo for 6-12 mo (random Effects model). Meta-analysis of fi ve highquality
trials including 693 women with low bone mass, revealed that a daily average of 56 mg soy Isoflavones for 6-12 mo tended to protect femoral neck BMD
compared with placebo (random effects model), the weighted mean differences
were 13.67 mg/cm2 (95% CI: -2.69 to 30.03; p =0.10), or 2.01% (95% CI: -0.39 to 4.40;
p =0.10). No significant Effects on hip total and trochanter BMD were found. When
the soy protein content labeling of 10 commercially available health foods (4 Foods
for Specifi ed Health Uses (FOSHU) and 6 health foods for an athlete) were assessed,
soy protein contents of one serving of these foods were almost the same as those
reported on the labels of the foods. Taken together with these findings and the results of the last year, there is possibility that intake of the foods fortifi ed with soy
protein or Isoflavones in addition to the normal diet improves bone density and fat
metabolism in postmenopausal women.
Soy Protein Research, Japan 12, 11-21, 2009.
Key words : extracted soy isoflavones, postmenopausal bone loss, bone mineral
density, health foods, soy protein
30-3
Transcriptome and Proteome Analyses of the Molecular Mechanism of a
Soybean Isoflavone, Genistein, in Overcoming Insulin Resistance
Kazumi YAGASAKI1, 2, Toshiaki SUEYASU3, Byung Geun HA2, Masato NAGAOKA1,
Takayuki YONEZAWA2 and Hisanori KATO3
1Institute of Symbiotic Science and Technology,
Tokyo University of Agriculture and Technology, Fuchu, Tokyo 183-8509
2Graduate School of Medicine, The University of Tokyo, Tokyo 113-0033
3Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo 113-8657
ABSTRACT
In a previous study, we found a soybean isoflavone genistein suppresses the
increase in the serum glucose level in type 2 diabetic model mice. To clarify
the mode of action of genistein, the effect of genistein on glucose metabolism
was investigated using rat L6 myotubes as a model of muscle tissue. Genistein
dose-dependently promoted glucose consumption in the absence of insulin in L6
myotubes cultured under normal and high glucose mediums. It is suggested that
by the results of examination using kinase inhibitors PI3-K, PKC and mTOR are
involved in the effect of genistein from the results of examination using various
kinase inhibitors. DNA microarray analysis revealed that 410 genes were upregulated
and 782 genes were down-regulated by the treatment with genistein.
The result of pathway analysis shows possibility of relation between the genistein's
Effects and signaling pathways of AMPK and IRS-1. Furthermore, we found that
genistein increased GLUT4 translocation into the plasma membrane and AMPK
phosphorylation.
Soy Protein Research, Japan 12, 22-26, 2009.
Key words : diabetes, genistein, muscle cells, glucose uptake, microarray
30-4
Identification of Genes Controlling the Contents of Seed Storage Proteins in Soybean
― Identification and Functional Analysis of the Quantitative Trait Locus qPro1 ―
Takanori YOSHIKAWA1, Shigeru UTSUMI2, Takako FUKUDA2, Yutaka OKUMOTO1,
Takashi SAYAMA1 and Takatoshi TANISAKA1
1Graduate School of Agriculture, Plant Breeding, Kyoto University,
2Graduate School of Agriculture, Food Quality Design and Development,
Kyoto University, Kyoto 606-8502
ABSTRACT
The contents of α'- and α-subunit of soybean β-conglycinin are controlled by
one major QTL, qPro1 , on linkage group (LG) I, which was detected using the
recombinant inbred lines derived from the cross between two varieties, Peking and
Tamahomare (PT-RILs). We first investigated the accumulation pattern of β-conglycinin subunits in Peking and Tamahomare during the seed ripening stage;
consequently, in both varieties, α'- and α-subunit showed similar accumulation
patterns, and the contents of them were both maximum at the 60% ripening stage.
This suggested the high possibility that qPro1 regulates the contents of these two
subunits at the expression level. The α-subunit content of a heterozygous RIL for
qPro1 was similar to that of Tamahomare; hence, Tamahomare-type allele at qPro1
was considered to be dominant to Peking-type allele. The candidate region of qPro1
was narrowed down within the 6.8 cM on LG I. Two α-subunit coding genes, CG-2
and CG-3, were included within the candidate region, and sequence analysis of these
genes disclosed the 6.0 kb deletion including CG-3 in Peking. The RILs harboring
CG-3 all showed high α-subunit contents, whereas those without it all sowed lowα-subunit contents. These results indicate that qPro1 is CG-3 gene, and its Pekingtype
allele is not capable of producing α-subunit due to the lack of CG-3.
Soy Protein
Research, Japan 12, 27-32, 2009.
Key words : soybean, seed storage protein, QTL, α-subunit, CG-3
30-5
Modifi cation of Soybean Seed Components by Gene Silencing
Akira KANAZAWA, Atsushi NAGAMATSU and Yusuke IMOTO
Research Faculty of Agriculture, Hokkaido University, Sapporo 060-8589
ABSTRACT
There is an increasing need for developing a system suitable for analyzing gene
function in soybean because of the rapid accumulation of the information of the
genome. RNA silencing is a high-throughput tool for suppressing gene expression
in a sequence-specific manner, either through RNA degradation or through
transcriptional repression. We have developed a system of RNA silencing using
a virus vector, which is termed virus-induced gene silencing (VIGS), in soybean.
Here we examined whether changes in seed components can be induced by VIGS
of flavonoid 3'-hydroxylase (F3'H) and chalcone synthase (CHS) in soybean. After
confirming the induction of VIGS in plants inoculated with viruses carrying a
portion of F3'H or CHS , isoflavone content in seeds produced on the plants was
analyzed by HPLC. As a result, a decrease in isoflavone content in seeds was
detected in plants in which CHS VIGS was induced, although no significant change
was detected in plants in which F3'H VIGS was induced. The successful induction
of changes in seed components by CHS VIGS suggest that our VIGS system is
useful for the modification of seed components in soybean, which allows rapid
functional analysis of genes involved in the biosynthesis of seed components as well
as prediction of changes in seed components as a consequence of conventional or
molecular breeding.
Soy Protein Research, Japan 12, 33-39, 2009.
Key words : flavonoid biosynthesis, gene expression control, reverse genetics, RNA
silencing, soybean
30-6
Manipulation of Biosynthesis of Seed Metabolic Components in Soybean
by Gene Silencing
Masao ISHIMOTO, Kyoko TAKAGI and Keito NISHIZAWA
National Agricultural Research Center for Hokkaido Region, Sapporo 062-8555
ABSTRACT
Soybean [Glycine max (L.) Merrill] is one of the world's most economically important
crops. Elucidation of molecular, biochemical and physiological mechanisms of the
biosynthesis of the seed metabolic components provides valuable information for
improvement of seed quality. In this study, we evaluated the application of RNA
interference (RNAi) to study on biosynthetic mechanisms of seed components. First
we validated our RNAi vector to regulate metabolic compounds by the introduction
to somatic embryos. Expression of the gene for phytoene desaturase, a key enzyme
in carotenoid synthesis, was suppressed by RNAi in somatic embryos, resulting in
reduced levels of carotenoids and a characteristic photobleaching phenotype. Next,
two RNAi vectors containing distinct regions of β-amyrin synthase gene, which
encodes a key enzyme in the biosynthesis of saponins, were introduced to the
somatic embryos. The transgenic plants regenerated and reproduced seeds without
any phenotypic abnormality. The transgenic seeds exhibited the drastic reduction
of saponin contents by the suppression of the β-amyrin synthase genes. Our
results thus suggest that RNAi would be a powerful tool in the functional analysis
of the seed components in soybean with duplicated genome structure.
Soy Protein
Research, Japan 12, 40-45, 2009.
Key words : carotenoid, gene silencing, RNA interference, saponin, secondary
metabolite, seed component
30-7
Characterization of Candidate Genes Involved in Catabolism of Lutein in Soybean
Seeds and Metabolic Engineering for Generation of Lutein Rich Soybean
Kyohei KANAMARU, Takehiko HAYASE,
Tetsuya YAMADA and Keisuke KITAMURA
Graduate School of Agriculture, Hokkaido University, Sapporo 060-8589
ABSTRACT
In this study, we examined the relationships between high lutein trait of wild
soybean seeds and the expression levels of two genes encoding CCDs, which are
involved in lutein catabolism. Putative two genes, GmCCD1 and GmCCD4 were
selected from the soybean genome database based on the homology with CCD
genes in Arabidopsis and Chrysanthemum. Quantitative RT-PCR analysis revealed
that two genes were always expressed during seed development. The expression
levels of GmCCD genes of RIL with low lutein were slightly higher than that of RIL
with high lutein in the middle stage of seed development. Linkage analysis using
CAPS marker for GmCCD4 gene revealed that the GmCCD4 gene was located
on the linkage group D1a. However, the position was not correspondence to QTL
for lutein content detected on the linkage group D1a. In addition, we attempted to
generate transgenic soybean plants for stable accumulation of lutein in soybean
seeds by the metabolic engineering. We constructed RNAi vectors to knockdown
the expression of GmCCD1 or GmCCD4 gene under the control of cotyledon
specific promoter. The transformation of a Japanese soybean variety has being
proceeded by Agrobacterium-mediated method.
Soy Protein Research, Japan 12, 46-50, 2009.
Key words : Glycine max, Glycine soja, lutein, carotenoid cleavage dioxygenase,
RNAi, 11S globulin promoter
30-8
Verifying the Dyanamism of Protein Anabolism and
Catabolism in β-conglycinin-rich, Glycinin-deficient Species of Soybean
Tomiko ASAKURA, Tomoko TAMURA and Kaede TERAUCHI
Graduate School of Agricultural and Life Sciences The University of Tokyo, Tokyo 113-8657
ABSTRACT
Comprehensive analyses on gene expression profiles in two soybean species,
Tozan 205 as a glycinin-lacking cultivar and Tamahomare as its parental species,
were conducted using DNA microarray technique. The analysis was focused on
four different stages of seed maturing such as small pods and maturing seeds
with sizes of 2 mm, 5 mm and 12-13 mm. In both Tozan 205 and Tamahomare,
the gene expression profiles varied depending on maturing; in early maturing
stage, transcription factors such as WRKY and MYB were expressed more in
Tamahomare than in Tozan 205. In the final "edamame" stage, seed maturation
proteins; 35 KDa, PM30, PM22, as well as glutathione S-transferase (GST)
and ascorbate peroxidase (APX) were expressed more in Tozan 205 than in
Tamahomare. Throughout the maturity significantly higher expressions of GST and
APX were found in Tozan 205 (11S deficient species) than in Tamahomare. These
results suggest that a molecular event of glycinin deficiency affects glutathione
metabolism in which both of these two genes are involved.
Soy Protein Research,
Japan 12, 51-57, 2009
Key words : DNA microarray glycinin, seed maturation
30-9
Relationship Between Gel Structure and Molecular Attraction of Glycinin Gel
Takao NAGANO
Department of Clinical Nutrition, Kawasaki University of Medical Welfare,
Kurashiki 701-0193
ABSTRACT
Heat-induced gels of glycinin and soybean protein isolate (SPI) were prepared using
magnesium chloride as a coagulant. Viscoelastic properties and microstructures of
glycinin and SPI gels were investigated using dynamic viscoelastic measurements
(DVM) and confocal laser scanning microscopy (CLSM). DVM indicated that G'
of a glycinin gel is two times higher than that of a SPI gel at appropriate NaCl
concentrations. This difference of G' between two these gels was reduced by adding
2-mercaptoethanol (2-ME). The microstructures of glycinin and SPI gels using
CLSM were analyzed by two parameters: the fractal dimension and the average
density of network structures. The fractal dimension of glycinin gels increased
and the average density of their network structures decreased with increasing of
adding 2-ME concentration. These results suggested that S-S bonds are important
for the formation of glycinin gels and make the density of their network structures
high.
Soy Protein Research, Japan 12, 58-62, 2009.
Key words : glycinin, S-S bond, heat-induced gel, dynamic viscoelastic measurement,
confocal laser scanning microscopy
30-10
Analysis of Ca2+-induced Co-gelation of Soy Protein Isolate and Gellan Gum
Takashi NAKAMURA and Kouichiro WATABE
Food Engineering Laboratory Department of Agricultural Chemistry School of Agriculture
Meiji University, Kawasaki 214-8571
ABSTRACT
The Ca2+-induced co-gelation of Soy Protein Isolate (SPI) and Gellan Gum (Gellan) by
heating-cooling process was investigated. The structural and mechanical properties
of gels were analyzed by rheological instruments and some kind of microscope. In
a large deformation test, the stress rupture strength of the co-gel was higher than
that of the gel formed from SPI alone or Gellan alone. The fracture strain ratio of
the co-gel was lower than that of the Gellan-gel. The structural features of gels at
different length scales by using confocal laser scanning microscopy (CLSM) and
scanning electron microscopy (SEM) were observed. Results obtained with CLSM,
at the macro-scale, indicated the bicontinuous SPI and Gellan phases of the co-gel.
Results obtained with SEM, at the meso-scale, indicated the interaction between
SPI aggregates and Gellan strands at the phase boundaries of bicontinuous SPI/
Gellan. The structural and viscoelastic properties of the coexistence solution of SPI
and Gellan by heating-cooling in the gelling process were analyzed. After the fi rst
heating, the phase separation to SPI-rich and Gellan-rich occurred already. At low
temperature with cooling, the separated Gellan phase was coarser. The viscosity
of the coexistence solution after Ca2+-addition increased over the sum of SPI alone
and Gellan alone. The Ca2+-induced co-gelation of SPI and Gellan induces the phase
separation process that leads to the bicontinuous structure formation at both macro
and meso level. Consequently, the co-gel shows novel properties which are different
from the single component gel.
Soy Protein Research, Japan 12, 63-69, 2009.
Key words :
30-11
Coagulation Inhibition of Soy Milk with Nigari by Soybean
Water-Soluble Polysaccharides
Hidefumi YOSHII and Takeshi FURUTA
Graduate school of Engineering, Tottori University, Tottori 680-8552
ABSTRACT
Coagulation of soy milk is the most important step in the tofu-making process. The
amount of coagulant added into soy milk is an especially critical variable during
soy milk coagulation and other additives such as polysaccharides also affect the
coagulation rate. The main objective of this study was to confi rm the infl uence of
addition of soybean water-soluble polysaccharides (SSPS) on the coagulation rate of
soy milk with nigari. The protein content in soy milk was 50 g/L. The coagulation
rate was measured in terms of the weight ratio of the coagulated portion to the
uncoagulated portion. The eff ect of SSPS addition (0-15 wt%) to nigari solution was
investigated. The reaction processes of coagulation of soy milk induced by nigari
with and without SSPS were investigated. The SSPS addition to nigari solution
retarded the coagulation rates of soy milk, particularly at low temperatures
between 20 to 40℃. The coagulation of soy milk with nigari might be assumed as
a sequential two-step reaction, which comprised the polymerization reaction of soy
protein and gelation reaction of polymerized soy proteins. The gelation reaction of
polymerized soy proteins was well correlated with the Avrami equation. The SSPS
addition affected the activation energy of the rate constant of gelation reaction.
Soy
Protein Research, Japan 12, 70-74, 2009
Key words : tofu, nigari, coagulation, kinetics, ssps
30-12
Development of a Method of Eliminating Soy Bean Smell with
Lactic Acid Bacteria
Hideyuki SUZUKI and Kanoko HAYASHIDA
Graduate School of Science and Technology, Kyoto Institute of Technology, Kyoto 606-8585
ABSTRACT
The causative substance for the soybean smell, n-Hexanol, is a major problem
when soy products are used in the food industry. In this study, we showed that a
lactic acid bacterium, Lactobacillus brevis , can be used to reduce n-hexanal in the
medium.
Soy Protein Research, Japan 12, 75-77, 2009.
Key words : n-hexanal, aldehyde dehydrogenase, lactic acid bacteria
30-13
Characterization of Yeast Cell Surface Displayed Aspergillus Oryzae
β-Glucosidase 1 High Hydrolytic Activity for Soybean Isoflavone
Chiaki OGINO
Department of Chemical Science and Engineering,
Graduate School of Engineering, Kobe University
ABSTRACT
For efficient production of isoflavone aglycones from soybean isoflavones, we
isolated three novel types of β-glucosidase (BGL1, BGL3 and BGL5) from the
fi lamentous fungi Aspergillus oryzae . Three enzymes were independently displayed
on the cell surface of a yeast Saccharomyces cerevisiae as a fusion protein withα-agglutinin. Three β-glucosidase-displaying yeast strains hydrolyzed isoflavone
glycosides efficiently, but exhibited different substrate specificities. Among theseβ-glucosidases, BGL1 exhibited the highest activity and also broad substrate
specificity to isoflavone glycosides. Although glucose released from isoflavone
glycosides are generally known to inhibit β-glucosidase, the residual ratio of
isoflavone glycosides generated using BGL1-displaying yeast strain reached
approximately 6.2%, and the glucose concentration in reaction mixture was
maintained at lower level. This result indicated that the BGL1-displaying yeast
strain assimilated the glucose before they inhibited the hydrolysis reaction, and
effi cient production of Isoflavone aglycones was achieved by engineered yeast cells
displaying β-glucosidase. In addition, the optimal pH and temperature for BGL1
displayed on the cell surfaces of the yeast were 5.0 and 55℃, while the optima for
BGL1 secreted by the yeast were 4.0 and 55℃. The displayed BGL1 was stable
at higher pH compared with the secreted BGL1. In addition, the thermostability
of BGL1 was improved by displaying the enzyme on the yeast cell surfaces. In
addition, the displayed and secreted forms of BGL1 had similar substrate specificity.β-glucosidase hydrolyzes daidzin and genistin, which are the glycoside forms of soybean Isoflavones, to the aglycones. Isoflavone aglycones were effi ciently produced
by BGL1-displaying yeast from an Isoflavone mixture; at optimal temperature and
pH the rate of aglycone production was at least 15.8 g/l/h. After 144 h of reaction,
almost Isoflavones were converted to its aglycone by BGL1-displaying yeast. The
results of the present study demonstrate that BGL1-displaying yeast strains are
effective whole cell biocatalysts of isoflavone aglycone production.
Soy Protein
Research, Japan 12, 78-83, 2009.
Key words : β-glucosidase, isoflavone, Saccharomyces cerevisiae, Aspergillus
oryzae , cell surface display
30-14
Improvement of Yeast Stress Tolerance by Soy Peptides
-Study on the Effects of Soy Peptides on Yeast Lipid Metabolism-
Shingo IZAWA
Department of Applied Biology, Graduate School of Science and Technology,
Kyoto Institute of Technology, Kyoto 606-8585
ABSTRACT
We have previously reported that the cultivation of yeast cells in media containing
soy peptides can improve tolerance to freeze-thaw stress, indicating that soy
peptides are suitable ingredients of culture media to provide high-quality yeast
cells for frozen-dough technology. We further investigated the mechanisms of
the improved tolerance to freeze-thaw stress by soy peptides, and found that
soy peptides affect the size and numbers of intracellular organelles. Compared
with yeast cells cultured with casein peptone, cells cultured with soy peptides
showed less of intracellular lipid bodies, in which neutral lipids are accumulated.
The cultivation with soy peptides resulted in decreased levels of mRNA of lipid
metabolism related genes such as DGA1 and ERG6 . These results suggest that the
low-fat yeast cells can be supplied by the cultivation with soy peptides and can be
utilized as a dietary supplement.
Soy Protein Research, Japan 12, 84-88, 2009.
Key words : soy peptides, baker's yeast, ergosterol, neutral lipids, lipid particle
30-15
Effects of Soy Peptide and Coexistent Food Components on the Metabolism of
Isoflavone by Fecal Flora
1Motoi TAMURA, 1Tadashi IWAMI, 1Sachiko HORI
1National Food Research Institute, Tsukuba 305-8642
ABSTRACT
We investigated the Effects of soy oligosaccharides on equol production in mice fed
a soy peptide-supplemented diet. We also investigated the effects of soy peptide
on in vitro incubation of daidzein with human fecal fl ora. Mice were administered
either soy oligosaccharides plus isoflavones (the SOI group) or isoflavones
(the control CI group) daily for 18 days before dissection. The plasma equol
concentration was higher in the group. In in vitro incubation of daidzein with fecal
fl ora of mice, equol concentration was slightly higher in the SOI group. Our data
suggest that the coadministration of soy peptide and soy oligosaccharides promotes
equol production or absorption in mice. When we investigated equol production in
in vitro incubation of daidzein with human fecal fl ora, the addition of soy peptide
HI-NUTE DH to the fecal suspension increased equol production in an equol
producer. However, equol non-producers did not become equol producers by the
addition of soy peptide HI-NUTE DH.
Soy Protein Research, Japan 12, 89-93, 2009.
Key words : soy peptides, daidzein, equol, mouse, soy oligosaccharides
30-16
Effects of Soy Protein Isolate on Cecal Fermentation and
Microbiota Patterns in Weaning Rats
Naomichi NISHIMURA, Kahori HAMADA and Tatsuro YAMAMOTO
Faculty of Health and Welfare Science, Nayoro City University, Hokkaido 096-8641
ABSTRACT
To clarify the hypothesis that the administration of different proteins after weaning
modifi es the intestinal micirobiota and fermentation, we examined the eff ect of soy
protein isolate (SPI) on cecal fermentation and microbiota patterns in weaning rats.
Female rats were fed a casein diet during gestation and lactation. After lactation for
21 d, litters were fed the casein and SPI diets for 3, 7, 14, 21 and 28 d in experiment
1, and for 7 and 21 d in experiment 2. In experiment 2, for the subsequent 7 and
14 d the diets were reversed. Cecal microbiota and cecal short chain fatty acid
concentration were determined. According to 16S rDNA PCR-DGGE analysis,
feeding the pups with SPI resulted in new clusters of cecal microbiota on at least
d 7 after weaning compared to the casein-fed pups. After 21 d post-weaning, the
microbiota patterns were similar. Cecal butyrate concentrations in the SPI-fed pups
were greater than in the casein-fed pups after 7 d post-weaning, and were stable
after 21 d post-weaning. In experiment 2, high cecal butyrate concentrations in SPIfed
pups decreased for 14 d after diet switching. In conclusion, when pups are fed
SPI as a protein source, cecal microbiota patterns different from those in caseinfed
pups are formed. The administration of SPI to weaning pups promotes cecal
butyrate production. However, this effect disappears if other protein is fed.
Soy
Protein Research, Japan 12, 94-99, 2009.
Key words : soy protein isolate, fermentation, microbiota, weaning, rats
30-17
Mechanism of Appetite Regulation by Daidzein.
―Approach Using Leptin- or Cholecystokinin-Receptor deficient-Animals―
Taro KISHIDA, Mina FUJITANI, Tomomi SEGAWA,
Asami IWAHARA and Kiyoshi EBIHARA
Department of Biological Resources, Faculty of Agriculture,
Ehime University, Matsuyama 790-8566
ABSTRACT
Previously, we have shown that soy isoflavone daidzein decreased food intake
specifically in female rats. Soy Isoflavones, especially equol, a metabolite of daidzein
by intestinal microorganisms, have estrogenic properties. Estrogen decreases food
intake, potentially by leptin or cholecystokinin-receptor (CCK)-related appetite
regulating system. We have aimed to confirm appetite regulation by daietary
daidzein, using leptin- and CCK-receptor deficient-rats, namely Zucker-obese rats
and OLETF rats. Contrary to the hypthesis, in the present study, we found that
dietary daidzein and estrogen treatment decreased food intake regardress of CCKreceptor
deficiency. CCK might not be a contributing factor in the mechanism
of daidzein and estrogen on decreased food intake in female rats. We also found
that estrogen treatment decreased food intake regardress of leptin-receptor
deficiency. Leptin might not contribute to the decreasing mechanism of estrogen
on food intake in female rats. Both Zucker-obese rat and Zucker-lean rats, the
origin of Zucker-obese rat, did not have enough equol production ability to decrease
food intake. Further experiment is needed to confirm the contribution of leptin
sensitivity to decreasing food intake by dietary daidzein.
Soy Protein Research, Japan 12, 100-108, 2009.
Key words : soy Isoflavone, daidzein, equol, leptin, CCK
30-18
Influence of Rat Strain and Diet Composition on Dietary
β-Conglycinin-Dependent Reduction of Food Consumption
Kazunori KOBA1, Daichi OIKAWA1, Asuka AKAHOSHI2, Sizuka TAMARU1,
Kazunari TANAKA1 and Michihiro SUGANO3
1Faculty of Nursing and Nutrition, University of Nagasaki, Siebold, Nagasaki 851-2195
2Faculty of Environmental and Symbiotic Sciences, Prefectural University of Kumamoto,
Kumamoto 862-8502
3Professor Emeritus, Kyushu University, Fukuoka 813-0043
ABSTRACT
We previously observed that the feeding of β-conglycinin (CON) as compared with
casein (CAS) and soy protein isolate (SPI) lowered body weight gain in rats, due
to a significant decrease in food consumption. In the present study, we examined
whether the CON-dependent decrease in food consumption was infl uenced by diet
composition (AIN-93G or AIN-76) and/or the rat strain (Sprague Dawley (SD) or
Wistar). Male SD or Wistar rats were fed diets prepared according to the AIN-
93G or AIN-76 formulas, containing 20% protein: CAS, SOY, CON; or a 1:1 mixture
of CAS and CON (CAS+CON). After a 4-week feeding period, dietary CON, as
compared with CAS and SPI, reduced food consumption and consequently body
weight gain, irrespective of diet formula and rat strain. The CON-dependent eff ect
was not observed at all in rats fed the CAS+CON diet. Therefore, it is suggested
that decreased food consumption resulting from the CON diet could be independent
on the diet formula and/or rat strain but is instead dependent on the protein
itself. Also, in rats fed the CAS+CON diets, CON-dependent physiological Effects
were still observed at a significant level, such as decreases in white adipose tissue
weights, serum triglyceride concentrations and hepatic fatty acid synthesis, and
increases in serum adiponectin concentration and fatty acid β-oxidation ability. The
results of the present study provide basic information for the elucidation of the physiological functions of CON.
Soy Protein Research, Japan 12, 109-114, 2009.
Key words : β-conglycinin, AIN-93G, rat strain, food intake, adipose tissue
30-19
The Mechanism of Dietary Soy Protein Regulation of Liver Lipid Concentration
Asako TAKENAKA
School of Agriculture, Meiji University, Kawasaki 214-8571
ABSTRACT
Effects of dietary soy protein isolate on hepatic fatty acid synthesis and on insulin
secretion were investigated. Male Wistar rats of 4 wk old were divided into two
groups and fed diets containing 20% casein (CAS) or soy protein isolate (SPI). After
two weeks, rats of each experimental group were sacrificed at three different
feeding conditions (fasted for 16 hours, refed for 2 h or 24 h after 16 h fasting).
Hepatic mRNA content of fatty acid synthase and acetyl-CoA carboxylase were
lower in SPI fed rats of fasted and 24 h refed groups. As for transcription factors
related to fatty acid synthesis, SRESBP-1c mRNA content was lower in SPI fed
rats of 2 h refed group. In order to elucidate the mechanism to reduce SREBP-
1c mRNA level, eff ect of SPI feeding on insulin secretion was investigated. Male
Wistar rats of 4 wk old were fed CAS or SPI diet for 14 d (9:00-17:00). On the day
3, 7 and 13 of the experiment, plasma samples were taken at 9:00, 10:00, 11:00 and
17:00 to measure insulin and glucose concentrations. The increase of plasma insulin
concentration after feeding was observed in rats fed CAS diet but was suppressed
in rats fed SPI diet for 3 d or more. These results demonstrate that even three
days feeding of SPI diet suppressed insulin secretion dramatically and that this may
also suppress SREBP-1 synthesis and contribute to reduction fatty acid synthesis in
liver.
Soy Protein Research, Japan 12, 115-119, 2009.
Key words : soy protein isolate, triglyceride, liver, rat, SREBP-1
30-20
Exaustive Analysis of Peptides Derived from Soybean Protein
with Bile Acid Binding Ability through Use of the Peptide Array
Satoshi NAGAOKA
Faculty of Applied Biological Sciences, Gifu University, Gifu 501-1193
ABSTRACT
This experiment was designed to evident peptides which have bile acid-binding
ability from soybean beta-conglycinin. In this study, we used a peptide array
to evaluate the bile acid binding ability of peptide derived from soybean betaconglycinin.
In the peptide array, bile acid binding ability was evaluated for the
binding ability of taurocholic acid and fixed peptides on cellulose menbrane,
then binding peptides with taurocholic acid were detected by first antibody of
bile acid, and finally the antibody-taurocholic acid complexes were detected by
second antibody. High bile acid-binding ability regions were evaluated in the fi rst
screening. In second screening, only peptides having stability with pepsin and
trypsin were designed and evaluated. We selected PVNKPG, IPFPRP, IPVNKPSR,
PRPIPF, RPIPFP, NVISQIPS, PIPFPR, PRPIPFPR to evaluate bile acid-binding
peptides with a digestive enzyme resistance. PVNKPG had a higher bile acidbinding
capacity in vitro than other synthesized peptides. IPFPRP or PVNKPG had
a lower micellar solubility of cholesterol than other synthesized peptides in vitro .
In Caco-2 cells models of small intestine, the uptake of cholesterol of PVNKPG had
the lowest value among the peptides groups. Thus, we found that PVNKPG
derived from soybean β-conglycinin bound taurocholate and inhibited cholesterol
absorption in Caco-2 cells.
Soy Protein Research, Japan 12, 120-124, 2009.
Key words : beta-conglycinin, peptide array, PVNKPG, cholesterol, bile acid
30-21
Screening of Soybean Isoflavones Inducing Gene Expression of
An Insulin-inducible Transcription Factor, SHARP-2
Kazuya YAMADA, Ayumi HANEISHI, Katsuhiro TAKAGI and Kosuke ASANO
Faculty of Human Health Science, Matsumoto University, Matsumoto 390-1295
ABSTRACT
The rat enhancer of split- and hairy-related protein-2 (SHARP-2) is an insulininducible
gene that encodes a transcriptional repressor. In the present study,
soybean constituents which induce expression of the SHARP-2 gene were screened.
Genistein, a soybean Isoflavone, induced the level of SHARP-2 mRNA in rat highlydifferentiated
H4IIE hepatoma cells. This induction was rapid and temporary,
and showed a dose-dependent manner. In contrast, Daidzein, another soybean
Isoflavone, did not induce the SHARP-2 gene expression. To examine whether an
induction of SHARP-2 gene expression by genistein as well as insulin is mediated
by a phosphoinositide 3-kinase (PI 3-K) pathway, H4IIE cells were treated with a PI
3-K inhibitor, LY294002. The induction by genistein was not inhibited by treatment
with LY294002. Thus, genistein specifically induced expression of the SHARP-2 gene
by a PI 3-K-indepepndent pathway.
Soy Protein Research, Japan 12, 125-128, 2009.
Key words : soybean Isoflavones, genistein, daidzein, transcription factor, SHARP-2,
insulin
30-22
Flavonoids Stimulate NGF-Induced Neurite Outgrowth in PC12 Cells
Yoshinori MARUNAKA, Naomi NIISATO,
Hiroaki MIYAZAKI and Ken-ichi NAKAJIMA
Graduate School of Medical Science,
Kyoto Prefectural University of Medicine, Kyoto 602-8566
ABSTRACT
During differentiation, neuronal cells undergo drastic shape changes. Typically,
neuronal cells extend long membranous processes, named neurites. PC12 cells
are derived from rat pheochromocytoma, and are a model useful for the study
on neuronal differentiation and neuro-regeneration because PC12 cells extend
neurites in response to nerve growth factor (NGF). We have recently reported that
Na+/K+/2Cl- cotransporter isoform 1 (NKCC1) plays an essential role in NGFinduced
neurite outgrowth of PC12D cells (a subclonal cell line of PC12). Flavonoids
are plant pigments widely contained in soybean, fruits and vegetables. Recent
studies have revealed that flavonoids modulate various cellular functions such as
cell cycle progression, apoptosis, and gene expression. In addition, our previous
studies have indicated that some flavonoids stimulate ion transport. Quercetin
and genistein stimulate transepithelial Cl- transport by activating NKCC1 and/or
cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel in renal
and airway epithelial cells. These reports suggest that some flavonoids are one of
possible activator of NKCC1. In the present study, we investigated the effect of
flavonoids (quercetin and genistein) on NGF-induced neurite outgrowth of PC12
cells. Quercetin stimulated the NGF-induced neurite outgrowth in a dose-dependent
mannar. Similarly, genistein also stimulated the neurite outgrowth, although the
effect of genistein was slightly weaker than that of quercetin. Knock-down of
NKCC1 by RNAi method abolished the stimulatory effect of these flavonoids.
Treatment with quercetin had no eff ect on the protein expression of NKCC1. These results indicate that flavonoids enhance the NGF-induced neurite outgrowth via the
activation of NKCC1 on the plasma membrane.
Soy Protein Research, Japan 12, 129-
133, 2009.
Key words : PC12, Neurite outgrowth, Quercetin, Genistein, Na+/K+/2Cl-cotransportertes
30-23
Effects of Genistein and Daidzein on the Anxiety-Like Behaviors in Mice.
Takayoshi MAMIYA
Graduate School of Pharmaceutical Sciences, Meijo University, Nagoya 468-8503
ABSTRACT
Previously, we have reported that mice fed food pellets containing soybean powder
for 4 weeks show anti-anxious Effects in mice. In the frontal cortex of those mice,
the contents of 3, 4-dihydroxyphenylacetic acid and homovanilic acid decreased,
whereas dopamine contents were not changed. In this study, we investigated
the effects of genistein and daidzein, major components of soybean, on anxietylike
behaviors in mice. When genistein (1 mg/kg) or daidzein (1 mg/kg) was
administered subcutaneously to ICR strain of male mice, the ratios of time spent
in the open arms and the entry to the open arms increased compared to vehicletreated
mice in the elevated plus-maze. Similar Effects of these reagents were found
in the light/dark box test. The injection of genistein or daidzein into the frontal
cortex failed to affect the extracellular dopamine level. These results suggest
that genistein and daidzein have anti-anxious Effects and dopaminergic system is
involved in these Effects at least in part.
Soy Protein Research, Japan 12, 134-137, 2009.
Key words : anti-anxious effects, genistein, daidzein, dopaminergic system
30-24
Study on the Induced Expression of LDL Receptor by Soy Isoflavones
Ryuichiro SATO
Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo 113-8657
ABSTRACT
Based on our previous finding that LDL receptor mRNA is quite unstable, in the
current study we investigated the molecular mechanism of the mRNA stability.
The human LDL receptor mRNA has a long 3' untranslated region (―2.5 kb) with 4
potential AU-rich elements (ARE) that are thought to be involved in rapid turnover
of various mRNA. The deletion analyses of the untranslated region revealed that
the fi rst ARE close to the stop codon is critical for the rapid turnover. We aimed
to identify cytoplasmic proteins that interact with this ARE and determine the
turnover rate of LDL receptor mRNA. We identified one of such ARE-binding
proteins and found that the protein is implicated in the LDL receptor mRNA
turnover.
Soy Protein Research, Japan 12, 138-141, 2009.
Key words : LDL receptor, AU-rich element, mRNA stability
30-25
Effect of Soy Isoflavones on the Antigen-specific Immune Responses in Mice
Tohru SAKAI, Mariko NAKAMOTO, Sakina FUROKU,
Yuko HIROTA and Emi SHUTO
Institution of Health Bioscience, The University of Tokushima Graduate School,
Tokushima 770-8503
ABSTRACT
The soy Isoflavone is a phytoestrogen found in high levels in soy products that has
been associated with decreased incidences of breast and prostate cancers. In this
study, we examined effect of daidzein and equol on the antigen-specific immune
responses in ovalbumin (OVA)-immunized BALB/c mice. Groups of mice were
exposed to the vehicle, daidzein or equol by gavage. The doses of soy Isoflavones
used were 4 and 20 mg/kg body/d. OVA-specific proliferative responses, IFN-γ and IL-4 productions were compared to those in control mice. OVA-specific IgE
productions were increased in 20 mg/kg equol administered mice. IL-13 production
in mice administered with 20 mg/kg equol was significantly increased. Therefore,
the mechanism for the elevation of the level of IgE by equol is responsible for the
increment of IL-13 production by T cells.
Soy Protein Research, Japan 12, 142-148,
2009.
Key words : soy Isoflavone, daidzein, equol, mice, IL-13
30-26
Soybean Lectin-Mediated Drug Delivery:
A Novel Method to Prevent Periodontal Diseases
Mayumi HIROSE1, 2, Toshiyuki MURAI3, Junji MORITA4, Atsushi NAKAGAWA1,
Mamoru SUZUKI1, Hiroto KAWASHIMA5 and Kimio NISHIMURA4
1Institute for Protein Research, Osaka University, Osaka 565-0871
2Graduate School of Agriculture Kyoto University, Kyoto 606-8502
3Osaka University Graduate School of Medicine, Suita 565-0871
4Department of Food Science and Nutrition Faculty of Human Life and Science, Doshisha
Women's College of Liberal Arts, Kyoto 602-0893
5School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka 422-8526
ABSTRACT
Dental plaque is composed of hundreds of different bacterial species within
an organized structure, and its accumulation on teeth is one of the causes of
periodontal diseases. Porphyromonas gingivalis is a predominant periodontal
pathogen that expresses a number of potential virulence factors involved in the
pathogenesis of periodontitis. FimA, a P. gingivalis fi mbriae protein, interacts withα5β1 integrin on gingival epithelial cells of the host, and promotes both bacterial
adhesion and invasion to the targeted sites. To date, it is difficult to completely
eradicate periodontitis, partly because the anti-bacterial drugs to this pathogen
cannot stay for a long time in the mouth. In the present study, we developed a
novel method that enables effi cient stay of a drug carrier on the mucosal surface of
gingival epithelial cells. We fi rst coated a resin with the soybean lectin, which binds
to O -linked glycans of mucins, to make the resin effi ciently bind to the surface of
gingival epithelial cells. We also coated the resin with a fibronectin or the RGD
peptide, together with the soybean lectin, in order to use the resin to block α5 β1 integrin-mediated adhesion of bacterial FimA protein to gingival cells. As a
result, the resin bound to the surface of gingival epithelial cells and blocked the binding of FimA effi ciently. The new method described here should be useful for
the development of an effective oral preventive medicine for the plaque.
Soy Protein
Research, Japan 12, 149-152, 2009.
Key words : soybean lectin, periodontal disease, Porphyromonas gingivalis, FimA,
integrin
30-27
Protective Effects of Dilinoleoylphosphatidylethanolamine on Endoplasmic
Reticurum Stress Induced Neuronal and Pancreatic β-cell Death
Kaoru NAGAI and Koki ITO
Interdisciplinary graduate school of Medicine and Engineering,
University of Yamanashi, Chuo-shi 409-3898
ABSTRACT
In many neurodegenerative diseases, neuronal cell death is caused by endoplasmic
reticulum (ER) stress. ER stress is mainly induced by the accumulation of unfolded
or misfolded proteins. We have reported that a phospholipid, dilinoleoylphosphatid
ylethanolamine (DLPE) protected neuronal cells from endoplasmic reticulum (ER)
stress induced cell death. In this study, we examined the protective mechanisms
of DLPE. First, we evaluated the effects of DLPE on up-regulation of an ER
chaperone Grp78 which prevents protein misfolding in the ER, and Chop which
was mainly reported to contribute to the cell death. ER stressors tunicamycin
(TM) and thapsigargin (TG) increased both Grp78 and Chop. DLPE stimulated
Grp78 expression in TM treated cells. However, unexpectedly, DLPE stimulated
Chop expression, as well. Recently, autophagy which is one of the intracellular
degradation mechanisms was reported to attenuate ER stress in neuronal cells.
And phosphatidylethanolamine (PE), such as DLPE is a component of autophagy.
Thus, DLPE might protect neuronal cells by stimulating autophagy. By western
blot analysis of LC3, we found that DLPE stimulated LC3 conversion which
indicates formation of autophagosome membrane. Our data suggests that DLPE
protected the neuronal cells by stimulating autophagy. Next, we studied if DLPE
is protective to other cells from ER stress. ER stress in pancreas was reported to
cause diabetes. Hence, we examined the protective eff ect of DLPE on ER stress
induced pancreatic β-cell line Min6 cell death. DLPE protected Min6 cells from
TM and TG toxicity. In addition, DLPE attenuated saturated fatty acid palmitate toxicity which is known to be a model of type-2 diabetes. Our data suggests that
consumption of phospholipids, such as DLPE, may reduce the risk of diabetes as
well as neurodegenerative diseases.
Soy Protein Research, Japan 12, 153-157, 2009.
Key words :Neurodegenerative disease, ER stress, autophagy phospholipid,
diabetes
30-28
Effect of Soy Bean β-conglycinin on Nonalcoholic Fatty Liver Disease
Tomomi YAMAZAKI
National Institute of Health and Nutrition, Tokyo 162-8636
ABSTRACT
The prevalence of obesity in Western societies has increased dramatically, due in
large part to high-sucrose and high-fat diets. The consequences of obesity include
the emerging epidemics of hepatic steatosis and nonalcoholic fatty liver disease
(NAFLD). Fatty liver has been thought to be benign. However, it is a precursor
of nonalcoholic steatohepatitis (NASH), which progresses to cirrhosis in up to
25% of patients. Soy protein in composed of the two major proteins, β-conglycinin
and glycinin. β-conglycinin is known to decrease plasma TG concentrations and
considered to have protective Effects against fatty liver. Diets high in sucrose or fat
can result in NAFLD. We analyzed the Effects of soybean β-conglycinin on NAFLD
induced by sucrose, saffl ower oil, and butter in ddY mice. Mice were fed a highstarch
diet plus 20% (w/w) sucrose in the drinking water, a high-saffl ower oil diet
or high-butter diet containing casein or β-conglycinin for 11 wk. As a control, mice
were fed a high-starch diet with drinking water. Mice supplemented with sucrose
or fed saffl ower oil or butter showed increased liver triglyceride content compared
with that of control mice. β-conglycinin completely prevented sucrose, safflower
oil or butter-induced NAFLD. Sucrose increased SREBP-1c and target mRNAs,
and β-conglycinin completely inhibited these increases. Butter increased PPARγ2 and target mRNA, and β-conglycinin completely inhibited these increases.β-conglycinin is effective in preventing NAFLD in those over-consuming sucrose
and fat.
Soy Protein Research, Japan 12, 158-161, 2009.
Key words : NAFLD, β-conglycinin, sucrose, fat
30-29
Effect and Signifi cance Intake of Beta-Conglucinin Fruit-Flavored Tablets on
Hyperlipidemia in Subjects with Renal Dysfunction
Fukuhiro UEDA1, Yoshinari MATUMOTO1, Yuka YAMAMOTO1, Yoshihiro NAKAMURA1,
Yuki YAMAMOTO1, Keiko YOSHIMI1, Kanae SANDA1, Makoto OTOSHI2,
Kentaro OTOSHI2, Aya MAEDA3, Aki HARANO3 and Yutaka HARANO3
1Hagoromo University of International Studies, Sakai 592-8344
2Otoshi Medical Clinic, Osaka 530-0017
3Harano hospital, Minoh 562-0005
ABSTRACT
The use of fenofibrate in subjects with hyperlipidemia and renal dysfunction is
not recommended. We have reported that β-conglycinin (β-CG),one of the major
components of soybean protein reduces blood triglyceride (TG) and LDL-cholesterol
(LDL-C). β-CG (6.3g/day) was administered to 14 patients for 3 mo, who showed
hyperlipidemia (TG≥150, LDL-C≥140 mg/dl) and renal dysfunction (Cr≥1.0 mg/dl,
or eGFR <60). TG decreased from 174±16 (SE) to 137±6 mg/dl, LDL-C from 112±5 to 100±7 mg/dl) significantly, and no change of HDL-cholesterol (HDL-C). 1/
Cr (1.00±0.10 vs 1.10±0.10) and eGFR (46±4 vs 50±5 mL/min/1.73m2) increased
significantly after the administration of β-CG. β-CG seemed to be effective in the
reduction of TG as well as LDL-C with the possible improvement of renal function.
Soy Protein Research, Japan 12, 162-165, 2009.
Key words :β-conglycinin, soy-bean protein, dyslipidemia, chronic kidney disease,
renal dysfunction
Soy Protein Research (Japan)