U.S. patent application number 12/919344 was filed with the patent office on 2011-01-06 for inhibitor for blood phosphorus level elevation.
Invention is credited to Masanori Asada, Tadashi Kanaya, Tetsuya Ogawa, Mikiko Shimada, Yumi Uehara.
Application Number | 20110002902 12/919344 |
Document ID | / |
Family ID | 41090788 |
Filed Date | 2011-01-06 |
United States Patent
Application |
20110002902 |
Kind Code |
A1 |
Asada; Masanori ; et
al. |
January 6, 2011 |
INHIBITOR FOR BLOOD PHOSPHORUS LEVEL ELEVATION
Abstract
The present invention provides an inhibitor for blood phosphorus
level elevation comprising a lactic acid bacterium as an active
ingredient. The inhibitor for blood phosphorus level elevation of
the present invention is highly safe, is readily administerable,
and can sufficiently inhibit a blood phosphorus level
elevation.
Inventors: |
Asada; Masanori; (Osaka,
JP) ; Kanaya; Tadashi; (Osaka, JP) ; Ogawa;
Tetsuya; (Tokyo, JP) ; Shimada; Mikiko;
(Gunma, JP) ; Uehara; Yumi; (Gunma, JP) |
Correspondence
Address: |
RENNER OTTO BOISSELLE & SKLAR, LLP
1621 EUCLID AVENUE, NINETEENTH FLOOR
CLEVELAND
OH
44115
US
|
Family ID: |
41090788 |
Appl. No.: |
12/919344 |
Filed: |
March 2, 2009 |
PCT Filed: |
March 2, 2009 |
PCT NO: |
PCT/JP2009/053837 |
371 Date: |
August 25, 2010 |
Current U.S.
Class: |
424/93.45 ;
424/93.4; 435/252.1; 435/252.9 |
Current CPC
Class: |
A61K 9/4875 20130101;
A61P 3/12 20180101; A61P 3/00 20180101; A61P 13/12 20180101; A61K
35/74 20130101; A61P 31/12 20180101 |
Class at
Publication: |
424/93.45 ;
435/252.9; 435/252.1; 424/93.4 |
International
Class: |
A61K 35/74 20060101
A61K035/74; C12N 1/20 20060101 C12N001/20; A61P 3/00 20060101
A61P003/00 |
Foreign Application Data
Date |
Code |
Application Number |
Mar 19, 2008 |
JP |
2008-072218 |
Claims
1. An inhibitor for blood phosphorus level elevation comprising a
lactic acid bacterium as an active ingredient, wherein the lactic
acid bacterium is at least one selected from the group consisting
of Bifidobacterium bifidum, B. longum, B. infantis, B. animalis, B.
pseudolongum, B. dentium, Lactobacillus acidophilus, L. animalis,
L. brevis, L. bulgaricus, L. casei, L. delbrueckii, L. plantarum,
Lactococcus lactis subsp. lactis, Enterococcus faecium, and
Enterococcus faecalis.
2-3. (canceled)
4. The inhibitor for blood phosphorus level elevation according to
claim 1, wherein the inhibitor for blood phosphorus level elevation
is in an oral dosage form.
5. The inhibitor for blood phosphorus level elevation according to
claim 4, wherein the inhibitor for blood phosphorus level elevation
is in at least one form selected from the group consisting of soft
capsules, hard capsules, and seamless capsules.
6. The inhibitor for blood phosphorus level elevation according to
claim 4, wherein the inhibitor for blood phosphorus level elevation
has acid resistance.
7. The inhibitor for blood phosphorus level elevation according to
claim 4, wherein the inhibitor for blood phosphorus level elevation
is enteric.
8. The inhibitor for blood phosphorus level elevation according to
claim 1, further comprising an oligosaccharide.
9. The inhibitor for blood phosphorus level elevation according to
claim 5, wherein the inhibitor for blood phosphorus level elevation
has acid resistance.
10. The inhibitor for blood phosphorus level elevation according to
claim 5, wherein the inhibitor for blood phosphorus level elevation
is enteric.
11. The inhibitor for blood phosphorus level elevation according to
claim 4, further comprising an oligosaccharide.
12. The inhibitor for blood phosphorus level elevation according to
claim 5, further comprising an oligosaccharide.
13. The inhibitor for blood phosphorus level elevation according to
claim 6, further comprising an oligosaccharide.
14. The inhibitor for blood phosphorus level elevation according to
claim 7, further comprising an oligosaccharide.
15. The inhibitor for blood phosphorus level elevation according to
claim 8, further comprising an oligosaccharide.
16. The inhibitor for blood phosphorus level elevation according to
claim 4, further comprising an oligosaccharide.
Description
TECHNICAL FIELD
[0001] The present invention relates to an inhibitor for blood
phosphorus level elevation. More specifically, the present
invention relates to an inhibitor for blood phosphorus level
elevation effective in the prophylaxis and amelioration of
hyperphosphatemia associated with reduced kidney function.
BACKGROUND ART
[0002] The kidney plays a very important role in the regulation of
phosphorus metabolism. Reduced kidney function would cause
hyperphosphatemia due to the build-up of phosphorus in the body.
Generally, the blood phosphorus level of a healthy individual is
about 2.5 to 4.7 mg/dL. When a blood phosphorus level exceeds 5.5
mg/dL, a treatment (such as, drug administration) is needed to
lower the blood phosphorus level.
[0003] In the excess phosphorus condition such as
hyperphosphatemia, the production and secretion of parathyroid
hormone (PTH) are increased, and the growth of accessory thyroid
cells is enhanced, thereby inducing secondary hyperparathyroidism.
It has been revealed that hyperphosphatemia is involved in the
development of renal failure, the onset of cardiovascular
complications (for example, phosphorus is bound with calcium in the
blood and deposited on the arterial walls, thereby causing
arteriosclerosis), and the like. It has been reported that
hyperphosphatemia is not only a cause of cardiovascular disorders
but also an exacerbating factor to reduce life expectancy for
dialysis patients and patients with compromised renal function.
[0004] Since a positive correlation is observed between an
elevation in PTH level and an increase in fractional phosphate
excretion rate, this effect has been believed to be mostly due to
increased PTH secretion. However, possible involvement of any
components other than PTH has been presented. It has not thoroughly
understood for the phosphorus metabolism. Therefore, there is
currently no developed, effective blood phosphorus level lowering
agent taking advantage of the function of phosphorus metabolism.
The removal of phosphorus from the blood relies on dialysis for
patients with renal failure who have lost their renal function.
[0005] Dialysis patients and patients with impaired renal function
often undergo severe dietary restrictions to reduce the intake of
phosphorus and the like. However, even with dietary restrictions,
they still intake phosphorus of about 1200 mg per day. Since at
most about 1000 mg of phosphorus can be removed in one dialysis
session, about 3000 mg of phosphorus can be removed in 3 dialysis
sessions weekly. Thus, the phosphorus intake is often
excessive.
[0006] Therefore, dialysis patients and patients with impaired
renal function often take a phosphate binder together with dietary
restrictions in order to prevent the elevation in blood phosphorus
levels. Examples of phosphate binders include those that can adsorb
physically phosphorus in the intestinal tract via oral ingestion,
including calcium-containing phosphate binders and calcium-free
phosphate binders.
[0007] However, calcium-containing phosphate binders would lead to
excessive intake of calcium to cause the calcium elevation in
blood, thereby increasing a risk of cardiovascular disorders. In
addition, a calcium-containing phosphate binder, when administered
in conjunction with activated vitamin D, cannot be administered in
a sufficient amount.
[0008] Examples of calcium-free phosphorus binders include
ion-exchange resin phosphate binders as disclosed in Patent
Documents 1 to 3. However, it is reported that there are side
effects such as constipation, abdominal fullness, nausea, and
vomiting in ion-exchange resin phosphate binders.
[0009] Furthermore, dialysis patients and patients with impaired
renal function may be restricted on the intake of not only
phosphorus but also potassium, salts, water, and the like. However,
sufficient restriction on the intake of phosphorus is difficult in
the regard of nutrient intake. Therefore, there is a demand for an
effective and safe pharmaceutical preparation.
[0010] Attempts have recently been made to develop a blood
phosphorus level lowering agent from food materials. Examples of
such blood phosphorus level lowering agents include a preparation
for hyperphosphatemia which contains as an effective ingredient a
galactomannan hydrolysate composed of neutral saccharides (Patent
Document 4), a blood phosphorus level lowering agent which contains
chitosan oligosaccharides as an effective ingredient (Patent
Document 5), and a phosphorus absorption inhibitor which contains a
red-alga extract as an effective ingredient (Patent Document
6).
[0011] The preparation for hyperphosphatemia which contains as an
effective ingredient a galactomannan hydrolysate as disclosed in
Patent Document 4 is usually ingested with food, but it seems that
the preparation is ingested with water when taken without food.
Since dialysis patients are subject to the restrictions on the
intake of water, the dosage form of allowing for ingestion without
water is preferable to ingest the preparation for
hyperphosphatemia.
[0012] The blood phosphorus level lowering agent which contains
chitosan oligosaccharides as an effective ingredient as disclosed
in Patent Document 5 contains oligosaccharide as the principal
component, and it would not cause deteriorated mineral absorption
in the large intestine. However, it is possible that such a blood
phosphorus level lowering agent would cause diarrhea, dehydration,
and the like.
[0013] The effective ingredient red alga of the phosphorus
absorption inhibitor as disclosed in Patent Document 6 can bind
strongly to potassium, calcium, iron, magnesium, sodium, and the
like to absorb such minerals, which is needed for dialysis
patients, thereby causing mineral deficiency. Since mineral level
would be varied on dialysis, the phosphorus absorption inhibitor
which can absorb minerals is required to be administered carefully
for dialysis patients.
[0014] It is known that an enteric preparation containing a lactic
acid bacterium is administered to reduce neutral fat, uremic
substances, and the like in blood for dialysis patients (Patent
Document 7). Patent Document 7 discloses that such a preparation
ameliorates arteriosclerosis, and ameliorates uremia associated
with chronic kidney failure by reducing indoxyl sulfate, phenol,
and like substances. However, Patent Document 7 does not make any
references to a blood phosphorus level.
[0015] Patent Document 8 discloses that a specific type of lactic
acid bacterium accumulates phosphorus in the microbial cell in the
form of polyphosphoric acid. However, any correlation is made clear
in Patent Document 8 between the accumulation of polyphosphoric
acid in the microbial cell and blood phosphorus level.
[0016] [Patent Document 1] Japanese Laid-Open Patent Publication
No. 2001-48791
[0017] [Patent Document 2] Japanese Laid-Open Patent Publication
No. 9-295941
[0018] [Patent Document 3] WO 01/068106
[0019] [Patent Document 4] Japanese Laid-Open Patent Publication
No. 2007-22992
[0020] [Patent Document 5] Japanese Laid-Open Patent Publication
No. 2000-344802
[0021] [Patent Document 6] Japanese Laid-Open Patent Publication
No. 2001-2581
[0022] [Patent Document 7] Japanese Laid-Open Patent Publication
No. 2004-277296
[0023] [Patent Document 8] Japanese Laid-Open Patent Publication
No. 2006-176450
DISCLOSURE OF INVENTION
Problems to be Solved by the Invention
[0024] It is an object of the present invention to provide an
inhibitor for blood phosphorus level elevation that is highly safe,
is readily administerable, and can inhibit a blood phosphorus level
elevation.
Means for Solving the Problems
[0025] The present invention provides an inhibitor for blood
phosphorus level elevation comprising a lactic acid bacterium as an
active ingredient.
[0026] In an embodiment, the lactic acid bacterium is at least one
selected from the group consisting of genera Bifidobacterium,
Lactobacillus, Lactococcus, and Enterococcus.
[0027] In an embodiment, the lactic acid bacterium is at least one
selected from the group consisting of Bifidobacterium bifidum, B.
longum, B. infantis, B. animalis, B. pseudolongum, B. dentium,
Lactobacillus acidophilus, L. animalis, L. brevis, L. bulgaricus,
L. casei, L. delbrueckii, L. plantarum, Lactococcus lactis subsp.
lactis, Enterococcus faecium, and Enterococcus faecalis.
[0028] In one embodiment, the inhibitor for blood phosphorus level
elevation is an oral dosage form.
[0029] In one embodiment, the inhibitor for blood phosphorus level
elevation is at least one form selected from the group consisting
of soft capsules, hard capsules, and seamless capsules.
[0030] In one embodiment, the inhibitor for blood phosphorus level
elevation has acid resistance.
[0031] In a further embodiment, the inhibitor for blood phosphorus
level elevation is enteric.
[0032] In a further embodiment, the inhibitor for blood phosphorus
level elevation further comprises an oligosaccharide.
EFFECTS OF INVENTION
[0033] The inhibitor for blood phosphorus level elevation of the
present invention is highly safe, is readily administerable, and
can inhibit a blood phosphorus level elevation for people such as
dialysis patients and patients with impaired renal function.
Moreover, the inhibitor for blood phosphorus level elevation of the
present invention, which contains a lactic acid bacterium, can
ameliorate constipation.
BRIEF DESCRIPTION OF DRAWINGS
[0034] FIG. 1 is a schematic cross-sectional illustration showing
the configuration of a three-layer seamless capsule
preparation.
BEST MODE FOR CARRYING OUT THE INVENTION
[0035] The inhibitor for blood phosphorus level elevation of the
present invention contains a lactic acid bacterium. The "inhibition
for blood phosphorus level elevation" refers to reducing a blood
phosphorus level, or inhibiting elevation or the rate of elevation
in a blood phosphorus level.
[0036] Lactic acid bacteria are known to improve the intestinal
environment and reduce decomposition products, such as ammonia,
indole, and phenol, in the intestines.
[0037] Examples of lactic acid bacteria include microorganisms
belonging to the genera Bifidobacterium, Lactobacillus,
Lactococcus, Pediococcus, Streptococcus, Enterococcus, Leuconostoc,
Tetragenococcus, Oenococcus, and Weissella. Such microorganisms may
be used singly or as a combination of two or more. Among such
microorganisms, it is preferable to use those belonging to the
genera Bifidobacterium and Lactobacillus.
[0038] Examples of microorganisms belonging to the genus
Bifidobacterium include Bifidobacterium bifidum, B. longum, B.
infantis, B. animalis, B. pseudolongum, B. dentium, B. angulatum,
B. asteroides, B. boum, B. catenulatum, B. choerinum, B.
coryneforme, B. cuniculi, B. gallicum, B. gallinarum, B. globosum,
B. indicum, B. magnum, B. merycicum, B. minimum, B. parvulorum, B.
pseudocatenulatum, B. pullorum, B. ruminale, B. ruminantium, B.
saeculare, B. subtile, B. suis, and B. thermophilum.
[0039] Among such microorganisms belonging to the genus
Bifidobacterium, it is preferable to use Bifidobacterium bifidum,
B. longum, B. infantis, B. animalis, B. pseudolongum, or B.
dentium.
[0040] Examples of microorganisms belonging to the genus
Lactobacillus include Lactobacillus acidophilus, L. amylovorus, L.
animalis, L. brevis, L. brevis subsp. gravesensis, L. buchneri, L.
bulgaricus, L. casei, L. casei subsp. casei, L. casei subsp.
plantarum, L. casei subsp. tolerans, L. cellobiosus, L. curvatus,
L. delbrueckii, L. delbrueckii subsp. bulgaricus, L. delbrueckii
subsp. delbrueckii, L. delbrueckii subsp. lactis, L. divergens, L.
fermentum, L. fructosus, L. gasseri, L. hilgardii, L. kefir, L.
leicnmannii, L. paracasei, L. paracasei subsp. paracasei, L.
pentosus, L. plantarum, L. reuteri, L. rhamnosus, L. sakei, L.
sakei subsp. sakei, L. sanfrancisco, L. vaccinostrcus, and
Lactobacillus sp.
[0041] Among such microorganisms belonging to the genus
Lactobacillus, it is preferable to use Lactobacillus acidophilus,
L. animalis, L. brevis, L. bulgaricus, L. casei, or L.
delbrueckii.
[0042] Examples of microorganisms belonging to the genus
Lactococcus include Lactococcus lactis, L. lactis subsp. hordniae,
L. lactis subsp. lactis, and L. raffinolactis.
[0043] Among such microorganisms belonging to the genus
Lactococcus, it is preferable to use Lactococcus lactis or L.
raffinolactis.
[0044] Examples of microorganisms belonging to the genus
Enterococcus include Enterococcus avium, E. casseliflavus, E.
cecorum, E. durans, E. faecalis, E. faecium, E. gallinarum, E.
hirae, E. malodoratus, E. mundtii, E. pseudoavium, E. raffinosus,
E. saccharolyticus, E. seriolicida, E. solitarius, and E.
villorum.
[0045] Among such microorganisms belonging to the genus
Enterococcus, it is preferable to use E. faecalis or E.
faecium.
[0046] The inhibitor for blood phosphorus level elevation of the
present invention contains a lactic acid bacterium such that the
lactic acid bacterium can be administered in a viable cell count of
preferably 5.times.10.sup.7 to 5.times.10.sup.10 and more
preferably 1.times.10.sup.9 to 1.times.10.sup.10 per day.
[0047] For example, when the inhibitor for blood phosphorus level
elevation of the present invention is in the form of a soft
capsule, hard capsule, or seamless capsule as described below, a
lactic acid bacterium is contained in a viable cell count of
preferably 2.times.10.sup.9 to 5.times.10.sup.9 per gram of a
capsule.
[0048] The inhibitor for blood phosphorus level elevation of the
present invention may further contain an oligosaccharide. The
oligosaccharide can assist the growth and the proliferation of the
lactic acid bacterium contained in the inhibitor for blood
phosphorus level elevation of the present invention.
[0049] There is no particular limitation on oligosaccharides for
use in the inhibitor for blood phosphorus level elevation of the
present invention, and examples include lactulose, raffinose,
fructooligosaccharides, galactooligosaccharides,
xylooligosaccharides, isomaltooligosaccharides, and
mannooligosaccharides.
[0050] When the inhibitor for blood phosphorus level elevation of
the present invention contains an oligosaccharide, the
oligosaccharide is contained in an amount of preferably 50 to 1000
mg and more preferably 100 to 500 mg per 2 billion cells of the
lactic acid bacterium contained in the inhibitor for blood
phosphorus level elevation of the present invention.
[0051] Furthermore, the inhibitor for blood phosphorus level
elevation of the present invention may contain, in addition to the
lactic acid bacterium and the oligosaccharide, other components
such as an excipient, an aroma chemical, and a solvent. Such
components include vitamins (such as vitamin A, vitamin B1, vitamin
B2, vitamin B6, vitamin B12, vitamin C, vitamin D, vitamin E,
pantothenic acid, folic acid, nicotinic acid, inositol, and
.beta.-carotene), amino acids (such as glycine, histidine,
isoleucine, and glutamic acid), nucleic acids (such as adenine,
guanine, thymine, cytosine, uracil, and like nucleobases;
adenosine, guanosine, cytidine, thymidine, uridine, and like
ribonucleosides, and monophosphate compounds, diphosphate
compounds, and triphosphate compounds thereof, deoxyadenosine,
deoxyguanosine, deoxycytidine, deoxythymidine, deoxyuridine, and
like deoxyribonucleosides, and monophosphate compounds, diphosphate
compounds, and triphosphate compounds thereof), minerals (such as
calcium, magnesium, iron, zinc, and copper), aliphatic acids (such
as .alpha.-linoleic acid, eicosapentaenoic acid (EPA),
docosahexaenoic acid (DHA), and evening primrose oil), octacosanol,
casein digests, water-soluble dietary fiber, insoluble dietary
fiber, saccharides (such as starch, cellulose, cornstarch, chitin,
chitosan, sucrose, lactose, maltose, and glucose), other useful
materials that are approved as food products, food additives, and
the like. Such other components may be used singly or as a
combination of two or more.
[0052] Among such examples, it is preferable to use vitamin C,
vitamin E, or .beta.-carotene. For example, vitamin C is contained
in an amount of preferably 10 to 500 mg and more preferably 50 to
200 mg per 2 billion cells of the lactic acid bacterium contained
in the inhibitor for blood phosphorus level elevation of the
present invention. Vitamin E is contained in an amount of
preferably 0.5 to 30 mg and more preferably 1.5 to 10 mg per 2
billion cells of the lactic acid bacterium contained in the
inhibitor for blood phosphorus level elevation of the present
invention. .beta.-Carotene is contained in an amount of preferably
0.5 to 20 mg and more preferably 1 to 5 mg per 2 billion cells of
the lactic acid bacterium contained in the inhibitor for blood
phosphorus level elevation of the present invention.
[0053] Since the inhibitor for blood phosphorus level elevation of
the present invention contains a lactic acid bacterium as an active
ingredient, it is preferable that the lactic acid
bacterium-containing inhibitor is administered orally. In this
case, the inhibitor should be designed so that the lactic acid
bacterium can travel through the stomach, reach the intestines, and
grow therein. Since the pH in the stomach is 1 to 3, most of the
lactic acid bacteria orally ingested would be killed by the very
low pH. Generally, it is said that no more than 1/10000 of the
amount administered of lactic acid bacteria can reach the
intestines while retaining the proliferating ability. Therefore,
the influence of gastric acid should be avoided as much as possible
so that the lactic acid bacterium in the inhibitor for blood
phosphorus level elevation of the present invention can viably
reach the human intestines and proliferate therein.
[0054] Therefore, it is preferable that the inhibitor for blood
phosphorus level elevation of the present invention is in the form
of an acid-resistant capsule preparation. The configuration, shape,
and the like of the capsule preparation are not particularly
limited insofar as the capsule shell is resistant to gastric acid.
Examples of the capsule preparation include soft capsules, hard
capsules, and seamless capsules. That is, a desirable configuration
is such that gastric acid does not penetrate into the capsule or
contact the lactic acid bacterium. The capsule shell may be a shell
that does not dissolve at pH 4 or less and preferably at pH 1 to 3.
There is also no particular limitation on the encapsulation
method.
[0055] It is further preferable that the inhibitor for blood
phosphorus level elevation of the present invention is in the form
of an enteric capsule preparation. That is, a preferable form is
such that the capsule isolates the capsule content lactic acid
bacterium from the outer environment by the shell of capsule in
gastric acid, or an acidic fluid having a pH of 1 to 3 and the
capsule opens or breaks so that the capsule content lactic acid
bacterium can contact with the fluid outside the capsule in the
intestinal fluid, or a weakly acidic (neutral) to weakly alkaline
fluid having a pH of 5 or greater. The capsule shell may not
necessarily dissolve insofar as the lactic acid bacterium in the
capsule can contact with the fluid outside the capsule. There is
also no particular limitation on the encapsulation method.
[0056] Seamless Capsule Preparation
[0057] The form of a capsule to impart gastric acid resistance is
preferably a seamless capsule. The "seamless capsule" is a type of
soft capsule and refers to a capsule of the form in which the
content is encapsulated by a seamless shell. It is possible that
the seamless capsule has a multilayer structure consisting of two
or more layers, and it is preferable that the seamless capsule has
a multilayer structure consisting of three or more layers.
[0058] Production of a three-layer seamless capsule preparation
shall be described below. FIG. 1 is a schematic cross-sectional
illustration of a three-layer seamless capsule preparation. This
three-layer structure is composed of an innermost layer, an
intermediate layer surrounding the innermost layer, and an external
layer surrounding the intermediate layer. The innermost layer is
composed of a lactic acid bacterium and a non-aqueous solvent or a
solid component to form a suspension or a mixture with the lactic
acid bacterium (hereinafter this component is referred to as an
innermost layer material).
[0059] There is no particular limitation on the innermost layer
material. Examples include various oils and fats, aliphatic acids,
saccharide fatty acid esters, aliphatic hydrocarbons, aromatic
hydrocarbons, linear ethers, higher fatty acid esters, higher
alcohols, and terpenes. More specifically, examples include, but
are not limited to, soybean oil, sesame oil, palm oil, corn oil,
cotton seed oil, coconut oil, rapeseed oil, cacao butter, beef
tallow, lard, horse oil, whale oil, hydrogenated oils and fats
thereof that have a melting point of 40.degree. C. or lower,
margarine, shortening, glycerin fatty acid esters, sucrose fatty
acid esters, peppermint oil, .alpha.-pinene, and D-limonene. Such
innermost layer materials may be used singly or as a combination of
two or more.
[0060] Materials for use in the intermediate layer are those that
are mentioned in connection with the innermost layer materials and
that have a melting point of 20 to 50.degree. C. but are different
from that used in the innermost layer. More preferably, materials
that are solid at ordinary temperatures are used. The intermediate
layer can serve to inhibit the penetration of fluids and oxygen,
preventing any contact with gastric acid. Selection of material can
be made according to, for example, the duration of the capsule
storage.
[0061] Examples of the materials of the external layer (the
outermost layer when there are 3 or more layers) include mixtures
of proteins and water-soluble polyhydric alcohols, mixtures of
proteins, water-soluble polyhydric alcohols, and polysaccharides,
and mixtures of polysaccharides and water-soluble polyhydric
alcohols. Examples of proteins include gelatin and collagen.
Examples of water-soluble polyhydric alcohols include sorbitol,
mannitol, glycerol, propylene glycol, and polyethylene glycol.
Examples of polysaccharides include agar, gellan gum, xanthan gum,
locust bean gum, pectin, alginates, carrageenan, gum arabic,
dextrin, modified dextrin, starch, modified starch, pullulan,
pectin, and carboxymethylcellulose salts. When pectin, alginates,
gellan gum, or carrageenan is used, an alkali metal salt, an
alkaline earth metal salt, or the like can be approproately
added.
[0062] Production of the above-described three-layer seamless
capsule preparation is performed according to a technique well
known to those skilled in the art, for example, a dripping method
that uses triple nozzles as described in the specification of
Japanese Patent No. 1398836. The capsule thus formed is then dried.
An example of drying is air drying at ordinary temperature. For
example, air drying at a temperature of 5 to 30.degree. C. is
commonly practiced. The drying time is preferably 2 to 12 hours.
Vacuum drying or freeze drying can be also performed.
[0063] To impart acid resistance to the capsule shell of the
seamless capsule preparation, an acid resistant external layer is
formed, or the shell (outermost layer) of the formed seamless
capsule is treated to attain acid resistance.
[0064] An example of a method of forming an acid resistant external
layer is a method in which pectin, alginates, gum arabic, or the
like is added in a proportion of 0.01 to 20 parts by weight and
preferably 0.1 to 10 parts by weight per 100 parts by weight of
gelatin, agar, carrageenan, or the like that has a gelling
ability.
[0065] Examples of a method of imparting acid resistance to the
shell (outermost layer) of the formed seamless capsule include a
cross-linking treatment performed on the external layer (outermost
layer) of the seamless capsule and a coating treatment performed on
the surface of the seamless capsule. It is preferable to perform
such treatments singly or as a combination.
[0066] When a protein-containing external layer is subjected to a
cross-linking treatment, the seamless capsule is first prepared and
thoroughly washed with water. The water-washed seamless capsule is
introduced into an aqueous solution that contains a cross-linking
agent to cross-link the surface of the external layer. For the
cross-linking agent, conventional cross-linking agents are usable,
and examples include formaldehyde, acetaldehyde, propionaldehyde,
glyoxal, glutaraldehyde, cinnamadehyde, vanillylaldehyde, acetone,
ethyl methyl ketone, ethylene oxide, propylene oxide, potassium
alum, and ammonium alum. Generally, the treatment of the external
layer is performed by introducing 1 part by weight of a seamless
capsule into 50 to 100 parts by weight of an aqueous solution
containing 0.1 to 2 w/v % and preferably 0.5 to 2 w/v % of a
cross-linking agent and stirring for 10 to 300 seconds. The amount
of cross-linking agent and the duration of action are varied
depending on the cross-linking agent. The surface of the external
layer after a cross-linking treatment is thoroughly washed with
water to remove the aqueous solution containing the cross-linking
agent, and then water contained in the external layer is dried.
[0067] For the above-described cross-linking treatment of a
protein-containing external layer, cross-linking may be achieved by
way of an enzymatic treatment using a transglutaminase. In this
case, the external layer is treated by introducing 1 part by weight
of the produced seamless capsule into 50 to 100 parts by weight of
an aqueous solution containing 0.1 to 10 w/v % and preferably 0.5
to 2 w/v % of an enzyme and stirring for 1 to 300 minutes, and
washing and drying are performed in the same manner as described
above.
[0068] If a coating treatment is performed, the produced wet
seamless capsule is dried and then coated according to a commonly
used procedure using shellac, ethylcellulose,
hydroxypropylmethylcellulose, hydroxypropylcellulose, polyvinyl
pyrrolidone, cellulose TC-5, a vinylpyrrolidone-vinyl acetate
copolymer, zein, ethylene wax, or the like as a base, and castor
oil, rapeseed oil, dibutyl phthalate, polyethylene glycol,
glycerol, stearic acid, a fatty acid ester, sorbitan palmitate,
polyoxyethylene stearate, acetylated monoglyceride, or the like as
a plasticizer.
[0069] Furthermore, imparting enteric properties to the capsule
protects the lactic acid bacterium present in the capsule from
acidic fluids (for example, gastric acid) and the like in the
stomach. Imparting enteric properties is performed according to a
method commonly employed by a skilled person to produce enteric
capsules. In addition, an enteric shell can be provided by use of a
mixture containing gelatin and pectin as a material of the external
layer of the seamless capsule.
[0070] The shape of the seamless capsule preparation may be
spherical. The average diameter of the seamless capsule is 0.3 to
10 mm and preferably 1.5 to 8 mm.
[0071] The seamless capsule preparation obtained in this manner can
be stored at room temperature for six months or longer while
maintaining the activity of the lactic acid bacterium, and can be
stored for a long period of time of one year or longer when stored
at 10.degree. C. or lower.
[0072] Soft Capsule Preparation
[0073] A soft capsule preparation, as with the seamless capsule
preparation, contains a suspension of a lactic acid bacterium in a
non-aqueous solvent as its content, enclosed by a shell sheet. The
material of the shell sheet is the same as the material of the
external layer of the seamless capsule.
[0074] The soft capsule preparation can be prepared according to a
known technique, for example, the method described in the
specification of Japanese Patent No. 2999535. For example, using a
rotary die, a shell sheet is passed through the die while injecting
and loading the content, and encapsulated by heating. The oil,
serving as a release agent, is removed from the resultant soft
capsule by washing with a polar solvent (such as methanol, ethanol,
propanol, or isopropanol) to release the lactic acid bacterium in
the intestines. Thereafter, as is the case with the seamless
capsule, a cross-linking treatment and a coating treatment are
performed in combination or either of the treatments is performed,
in order to attain acid resistance.
[0075] The soft capsule preparation may be spherical, spheroidal,
or rectangular parallelepiped. A soft capsule having a major axis
of 3 to 16 mm and a minor axis of 2 to 10 mm is preferable, and a
soft capsule having a major axis of 5 to 7 mm and a minor axis of 2
to 4 mm is more preferable.
[0076] The soft capsule preparation obtained in this manner can be
stored at room temperature for six months or longer while
maintaining the activity of the lactic acid bacterium, and can be
stored for a long period of time of one year or longer when stored
at 10.degree. C. or lower.
[0077] Hard Capsule Preparation
[0078] A hard capsule preparation can be produced by molding in
advance a capsule shell into a body and a cap, loading the content
into the capsule body, and then fitting the capsule cap thereto.
Examples of materials of the hard capsule include gelatin,
cellulose, pullulan, hydroxypropylmethylcellulose, carrageenan, and
cellulose derivatives. The hard capsule can be molded according to
a method commonly employed by a skilled person. The molded capsule
is also a commercially available. The content can be a thorough
mixture of a lactic acid bacterium and an excipient (such as
silicic anhydride, synthetic aluminum silicate, lactose,
cornstarch, crystalline cellulose), or a powder of dried cells of a
lactic acid bacterium. A coating can be provided after loading the
content into the capsule. The materials and methods described in
connection with the external layer of the seamless capsule are
applicable to this coating, thereby imparting acid resistance and
preferably collapsibility in the intestines (enteric properties).
This coating can also serve to seal the capsule shell to envelope
the content.
[0079] The hard capsule preparation obtained in this manner can be
stored at room temperature for six months or longer while
maintaining the activity of the lactic acid bacterium, and can be
stored for a long period of time of one year or longer when stored
at 10.degree. C. or lower.
[0080] The present invention shall be described in detail below by
way of examples and comparative examples, but the present invention
is not limited to the examples.
EXAMPLES
Example 1
[0081] The seamless capsule containing bifidobacterium was prepared
as described below using capsule production apparatus equipped with
triple concentric nozzles.
[0082] The cell powder of Bifidobacterium (Bifidobacterium longum
subspecies longum JCM1217) in an amount of 100 g was dispersed in
hydrogenated oil prepared by melting 400 g of hydrogenated palm
kernel oil having a melting point of 34.degree. C. This dispersion
was discharged from the inner nozzle of the triple concentric
nozzles, hydrogenated oil prepared by melting hydrogenated palm
kernel oil having a melting point of 43.degree. C. was discharged
from the intermediate nozzle disposed around the inner nozzle, and
a gelatin solution (a solution prepared by dissolving 600 g of
gelatin, 300 g of glycerol, and 100 g of pectin in 4 kg of purified
water) was discharged from the outermost nozzle simultaneously into
rapeseed oil flowing as a fluid carrier under cooling (15.degree.
C.) in order to prepare the three-layer seamless capsule containing
bifidobacterium, having a diameter of 2.0 mm (2 billion cells of
bifidobacterium per 0.2 g of seamless capsule).
[0083] 34 dialysis patients (hereinafter sometimes simply referred
to as patients) ingested 0.2 g of the bifidobacterium-containing
seamless capsule thus prepared daily over 4 weeks. The patients did
not take any other blood phosphorus level lowering agents. While
under dietary instructions by a dietitian, not all patients had the
same meals.
[0084] Blood was collected before the ingestion of the
bifidobacterium-containing seamless capsule and 1 week, 3 weeks,
and 4 weeks after the beginning of ingestion, and blood phosphorus
levels were measured according to the molybdenum blue method using
a Phospha C-Test Wako for inorganic phosphorus measurement
(manufactured by Wako Pure Chemical Industries, Ltd.).
[0085] Tables 1 and 2 show the results of measuring the blood
phosphorus level of each patient. In addition, Table 3 shows the
results of measuring the blood phosphorus levels of 11 patients who
did not take either the bifidobacterium-containing seamless
capsules or any other blood phosphorus level lowering agents
(control group).
TABLE-US-00001 TABLE 1 Blood phosphorus level (mg/dL) 1 week after
3 weeks after 4 weeks after Before the beginnig of the beginnig of
the beginnig of ingestion ingestion ingestion ingestion Patient 1
6.35 5.8 5.9 4.7 Patient 2 6.08 4.9 5.5 4.5 Patient 3 5.53 5.1 5.0
4.1 Patient 4 6.08 4.7 3.9 4.9 Patient 5 5.93 5.0 4.8 4.8 Patient 6
8.25 6.1 7.0 6.7 Patient 7 6.53 6.2 6.3 5.8 Patient 8 5.48 4.2 3.6
4.9 Patient 9 6.15 5.7 5.2 5.7 Patient 10 7.23 7.5 7.2 6.7 Patient
11 6.58 6.1 5.9 6.1 Patient 12 6.80 6.6 7.9 6.4 Patient 13 5.80 6.2
6.1 5.6 Patient 14 7.33 5.9 6.3 7.1 Patient 15 6.48 4.4 5.6 6.3
Patient 16 7.18 6.3 7.4 7.1 Patient 17 8.13 4.8 -- 8.1
TABLE-US-00002 TABLE 2 Blood phosphorus level (mg/dL) 1 week after
3 weeks after 4 weeks after Before the beginnig of the beginnig of
the beginnig of ingestion ingestion ingestion ingestion Patient 18
5.80 5.5 6.1 5.8 Patient 19 6.88 5.9 7.5 6.9 Patient 20 6.53 5.4
5.9 6.6 Patient 21 8.43 8.0 9.5 8.6 Patient 22 5.68 5.8 5.5 5.8
Patient 23 6.45 8.0 6.6 6.7 Patient 24 5.93 5.3 5.7 6.2 Patient 25
6.63 5.5 6.1 7.0 Patient 26 6.15 6.2 7.6 6.5 Patient 27 6.15 5.4
6.5 6.7 Patient 28 6.47 7.2 6.4 7.1 Patient 29 7.83 8.1 8.5 8.7
Patient 30 6.40 6.9 7.1 7.3 Patient 31 7.45 8.0 9.4 8.5 Patient 32
5.37 6.2 6.2 6.6 Patient 33 6.83 7.7 7.0 8.5 Patient 34 8.23 9.6
8.8 10.9
TABLE-US-00003 TABLE 3 Blood phosphorus level (mg/dL) 1 week after
3 weeks after 4 weeks after Before the beginnig of the beginnig of
the beginnig of ingestion ingestion ingestion ingestion Patient 35
8.25 8.4 9.0 9.5 Patient 36 6.12 5.9 5.9 7.4 Patient 37 6.96 6.7
7.4 7.5 Patient 38 7.33 7.9 7.9 8.0 Patient 39 7.35 7.1 7.0 8.5
Patient 40 5.97 6.2 6.1 6.3 Patient 41 6.20 6.7 6.6 6.9 Patient 42
7.69 8.2 8.4 10.7 Patient 43 6.85 6.6 6.9 8.3 Patient 44 7.94 7.7
7.6 9.8 Patient 45 7.92 8.0 8.7 9.2 Average 7.1 7.2 7.4 8.4 Average
increase in blood phosphorus level for 4 weeks: 1.3 mg/dL
[0086] As seen from the values indicating the blood phosphorus
levels before the ingestion and after 4 weeks of the beginning of
ingestion of the bifidobacterium-containing seamless capsules
presented in Tables 1 and 2, the average increase in blood
phosphorus level for the 34 patients was -0.04 mg/dL (i.e., the
average blood phosphorus level was reduced by 0.04 mg/dL).
[0087] As shown in Table 1, it can be understood that for the 17
patients, the blood phosphorus levels 4 weeks after the ingestion
of the bifidobacterium-containing seamless capsules were lowered
compared to their blood phosphorus levels before the ingestion of
the bifidobacterium-containing seamless capsules.
[0088] On the other hand, as shown in Table 2, for the remaining 17
patients, the blood phosphorus levels were not lowered by the
ingestion of the bifidobacterium-containing seamless capsule.
However, since the average blood phosphorus level was reduced by
0.04 mg/dL for the 34 patients, it can be understood that a blood
phosphorus level elevation was inhibited.
[0089] For the control group, the blood phosphorus levels after
treatment were significantly higher than the levels before
treatment (Paired T-test: p<0.01), although no significant
increase was observed for the treated group. Therefore, it can be
understood that the bifidobacterium-containing seamless capsule
obtained in Example 1 significantly inhibit a blood phosphorus
level elevation.
Example 2
[0090] The cell powder of Bifidobacterium (Bifidobacterium longum
subspecies longum JCM1217) in an amount of 50 g was suspended in
300 g of rapeseed oil to prepare the content fluid of soft capsule.
Next, 400 g of gelatin and 100 g of glycerol were added to 200 g of
distilled water and dissolved by stirring at 60.degree. C., and the
solution was molded into a sheet to obtain gelatin films. Next, the
gelatin films were fed between a pair of rotary cylindrical metal
molds while the content fluid was injected between the gelatin
films with a pump operating in concert with the molds in order to
prepare the soft capsule (2 billion cells of bifidobacterium per
soft capsule).
[0091] 30 dialysis patients ingested 1 capsule of the
bifidobacterium-containing soft capsule thus prepared daily over 4
weeks. The condition of ingestion was the same as that in Examples
1. The patients did not take any other blood phosphorus level
lowering agents. While under dietary instructions by a dietitian,
not all patients had the same meals.
[0092] Blood was collected before and 4 weeks after the beginning
of the ingestion of the bifidobacterium-containing soft capsules,
and blood phosphorus levels were measured. Table 4 shows the
results.
[0093] For 15 patients who did not take either the
bifidobacterium-containing soft capsules or any other blood
phosphorus level lowering agents (control group), the average blood
phosphorus level before treatment was about 7.10 mg/dL, and the
average blood phosphorus level four-week after treatment was about
8.30 mg/dL (for the control group, the average increase in blood
phosphorus level was 1.21 mg/dL).
TABLE-US-00004 TABLE 4 Blood phosphorus level (mg/dL) 4 weeks after
Before the beginnig ingestion of ingestion Patient 1 5.7 5.1
Patient 2 8.1 7.5 Patient 3 8.3 7.9 Patient 4 6.3 5.9 Patient 5 7.1
7.0 Patient 6 8.0 7.9 Patient 7 7.2 7.2 Patient 8 7.0 7.0 Patient 9
6.0 6.5 Patient 10 6.7 7.4 Patient 11 6.0 6.7 Patient 12 6.0 6.8
Patient 13 7.6 8.4 Patient 14 7.2 8.2 Patient 15 8.4 9.6 Patient 16
7.3 8.7 Patient 17 6.0 7.3 Patient 18 8.2 9.6 Patient 19 6.1 7.6
Patient 20 5.7 7.2 Patient 21 7.0 8.6 Patient 22 8.3 10.0 Patient
23 7.6 9.3 Patient 24 7.5 9.3 Patient 25 5.5 7.3 Patient 26 6.0 7.9
Patient 27 6.7 8.6 Patient 28 7.2 9.2 Patient 29 8.0 10.0 Patient
30 5.9 8.0 Rate of patients with reduced blood phosphorus level:
20% (6 patients) Average increase in blood phosphorus level for
patients 1 to 30: 0.97 mg/dL Average increase in blood phosphorus
level for control group: 1.21 mg/dL
[0094] As shown in Table 4, it can be understood that 6 (patients 1
to 6) of the 30 patients had lowered blood phosphorus levels after
the ingestion of the bifidobacterium-containing soft capsule. On
the other hand, lowering of the blood phosphorus level was not
observed after the ingestion of the bifidobacterium-containing soft
capsule for the remaining 24 patients. The average increase in
blood phosphorus level was 0.97 mg/dL for the 30 patients compared
to the increment for the control group of 1.21 mg/dL, and it can be
understood that a blood phosphorus level elevation was
inhibited.
Example 3
[0095] A coating was applied to the soft capsule as obtained in
Example 2 according to the method described in Japanese Laid-Open
Patent Publication No. 2003-230363.
[0096] First, an immersion fluid containing 20 parts by weight of
shellac, 2 parts by weight of triethyl citrate, and 78 parts by
weight of ethanol was prepared. Next, the soft capsule as obtained
in Example 2 was immersed in the immersion fluid thus prepared, and
subject to air-drying at about 15 to 20.degree. C. such that the
viable cell count of the bifidobacterium was not reduced. The
immersion and drying treatment was repeated 3 times, in order to
prepare the shellac-coated soft capsule containing bifidobacterium
(enteric bifidobacterium-containing soft capsule).
[0097] 30 dialysis patients ingested 1 capsule of the enteric
bifidobacterium-containing soft capsule thus prepared daily under
the same conditions of ingestion as in Example 1. Blood was
collected before and 4 weeks after the beginning of the ingestion
of the enteric bifidobacterium-containing soft capsule, and blood
phosphorus levels were measured. Table 5 shows the results.
[0098] For 15 patients who did not take either the enteric
bifidobacterium-containing soft capsules or any other blood
phosphorus level lowering agents (control group), the average blood
phosphorus level before treatment was about 6.86 mg/dL, and the
average blood phosphorus level four-week after treatment was about
8.02 mg/dL (for the control group, the average increase in blood
phosphorus level was 1.15 mg/dL).
TABLE-US-00005 TABLE 5 Blood phosphorus level (mg/dL) 4 weeks after
Before the beginnig ingestion of ingestion Patient 1 6.0 4.7
Patient 2 7.5 6.1 Patient 3 5.7 4.4 Patient 4 7.7 6.6 Patient 5 6.6
5.6 Patient 6 8.0 7.0 Patient 7 5.5 4.6 Patient 8 8.2 7.4 Patient 9
8.1 7.2 Patient 10 5.4 4.7 Patient 11 6.4 5.7 Patient 12 6.4 5.7
Patient 13 7.9 7.7 Patient 14 8.3 8.2 Patient 15 7.8 8.0 Patient 16
7.4 7.7 Patient 17 5.9 6.3 Patient 18 7.8 8.3 Patient 19 7.9 8.7
Patient 20 8.1 9.1 Patient 21 5.5 6.5 Patient 22 7.7 8.7 Patient 23
7.1 8.2 Patient 24 7.7 8.8 Patient 25 5.8 7.1 Patient 26 7.6 8.9
Patient 27 8.3 9.7 Patient 28 8.0 9.4 Patient 29 6.8 8.3 Patient 30
5.7 7.2 Rate of patients with reduced blood phosphorus level: 47%
(14 patients) Average increase in blood phosphorus level for
patients 1 to 30: 0.13 mg/dL Average increase in blood phosphorus
level for control group: 1.15 mg/dL
[0099] As shown in Table 5, it can be understood that 14 (patients
1 to 14) of the 30 patients had lowered blood phosphorus levels
after the ingestion of the enteric bifidobacterium-containing soft
capsule. On the other hand, lowering of the blood phosphorus level
was not observed after the ingestion of the
bifidobacterium-containing soft capsule for the remaining 16
patients. The average increase in blood phosphorus level was 0.13
mg/dL for the 30 patients compared to the increment for the control
group of 1.15 mg/dL, and it can be understood that a blood
phosphorus level elevation was inhibited.
[0100] For the control group, the blood phosphorus levels after
treatment were significantly higher than the levels before
treatment (Paired T-test: p<0.01), although no significant
increase was observed in the treated group. Therefore, it can be
understood that the enteric bifidobacterium-containing soft capsule
obtained in Example 3 significantly inhibit a blood phosphorus
level elevation.
Example 4
[0101] The cell powder of bifidobacterium (Bifidobacterium longum
subspecies longum JCM1217) was loaded into a commercial hard
capsule having a size according to the Japanese Pharmacopoeia of 5
in 2 billion cells of bifidobacterium per 1 hard capsule to prepare
the hard capsule containing bifidobacterium.
[0102] Next, 100 g of the hard capsule thus prepared was placed in
a tumbling granulator, and a solution prepared by dissolving 10 g
of shellac and 1 g of castor oil in 400 g of a mixture of methanol
and ethyl acetate (volume ratio 1:1) was sprayed onto the entire
surface of the hard capsule to a coating thickness of 0.3 mm, in
order to prepare 100 g of the hard capsule with acid resistant
coating (enteric bifidobacterium-containing hard capsule).
[0103] 30 dialysis patients ingested 1 capsule of the enteric
bifidobacterium-containing hard capsule thus prepared daily under
the same conditions of ingestion as in Example 1. Blood was
collected before and 4 weeks after the beginning of the ingestion
of the enteric bifidobacterium-containing hard capsule, and blood
phosphorus levels were measured. Table 6 shows the results.
[0104] For 15 patients who did not take either the enteric
bifidobacterium-containing soft capsules or any other blood
phosphorus level lowering agents (control group), the average blood
phosphorus level before treatment was about 7.02 mg/dL, and the
average blood phosphorus level four-week after treatment was about
8.01 mg/dL (for the control group, the average increase in blood
phosphorus level was 0.99 mg/dL).
TABLE-US-00006 TABLE 6 Blood phosphorus level (mg/dL) 4 weeks after
Before the beginnig ingestion of ingestion Patient 1 6.3 4.9
Patient 2 7.0 5.8 Patient 3 6.1 4.9 Patient 4 5.6 4.4 Patient 5 7.4
6.2 Patient 6 7.6 6.6 Patient 7 7.5 6.7 Patient 8 8.0 7.4 Patient 9
6.6 6.0 Patient 10 6.2 5.7 Patient 11 7.0 6.5 Patient 12 6.5 6.1
Patient 13 6.6 6.3 Patient 14 5.8 5.6 Patient 15 6.0 5.9 Patient 16
6.7 6.6 Patient 17 8.2 8.3 Patient 18 6.8 7.0 Patient 19 6.9 7.3
Patient 20 5.8 6.3 Patient 21 7.1 7.7 Patient 22 7.4 7.9 Patient 23
7.8 8.5 Patient 24 8.3 9.0 Patient 25 7.5 8.3 Patient 26 8.2 9.1
Patient 27 6.9 8.1 Patient 28 6.2 7.4 Patient 29 7.0 8.4 Patient 30
7.4 9.0 Rate of patients with reduced blood phosphorus level: 53%
(16 patients) Average increase in blood phosphorus level for
patients 1 to 30: 0.01 mg/dL Average increase in blood phosphorus
level for control group: 0.99 mg/dL
[0105] As shown in Table 6, it can be understood that 16 (patients
1 to 16) of the 30 patients had lowered blood phosphorus levels
after the ingestion of the enteric bifidobacterium-containing hard
capsule. On the other hand, lowering of the blood phosphorus level
was not observed after the ingestion of the enteric
bifidobacterium-containing hard capsule for the remaining 14
patients. The average increase in blood phosphorus level was 0.01
mg/dL for the 30 patients compared to the increment for the control
group of 0.99 mg/dL, and it can be understood that a blood
phosphorus level elevation was inhibited.
[0106] For the control group, the blood phosphorus levels after
treatment were significantly higher than the levels before
treatment (Paired T-test: p<0.01), although no significant
increase was observed in the treated group. Therefore, it can be
understood that the enteric bifidobacterium-containing hard capsule
obtained in Example 4 significantly inhibit a blood phosphorus
level elevation.
Example 5
[0107] In order to prepare the Bifidobacterium-containing seamless
capsule, the procedure was followed as in Example 1 except that the
powder of killed bifidobacterium (Bifidobacterium longum subspecies
longum JCM1217) cells was prepared by treating 100 g of the powder
of the viable cells at 500 W for 10 minutes in a microwave
oven.
[0108] 30 dialysis patients ingested 0.2 g of the dead
bifidobacterium-containing seamless capsule thus prepared daily
under the same conditions of ingestion as in Example 1. Blood was
collected before and 4 weeks after the beginning of the ingestion
of the dead bifidobacterium-containing seamless capsule, and blood
phosphorus levels were measured. Table 7 shows the results.
[0109] For 15 patients who did not take either the killed
bifidobacterium-containing seamless capsules or any other blood
phosphorus level lowering agents (control group), the average blood
phosphorus level before treatment was about 7.19 mg/dL, and the
average blood phosphorus level four-week after treatment was about
8.55 mg/dL (for the control group, the average increase in blood
phosphorus level was 1.36 mg/dL).
TABLE-US-00007 TABLE 7 Blood phosphorus level (mg/dL) 4 weeks after
Before the beginnig ingestion of ingestion Patient 1 6.6 6.4
Patient 2 5.6 5.4 Patient 3 7.7 7.5 Patient 4 6.2 6.1 Patient 5 7.9
7.9 Patient 6 6.3 6.4 Patient 7 7.8 7.9 Patient 8 6.5 6.6 Patient 9
7.9 8.1 Patient 10 8.3 8.6 Patient 11 7.7 8.0 Patient 12 8.4 8.7
Patient 13 6.5 6.9 Patient 14 6.9 7.4 Patient 15 7.4 8.4 Patient 16
5.7 6.7 Patient 17 7.5 8.7 Patient 18 8.1 9.3 Patient 19 5.9 7.3
Patient 20 6.8 8.3 Patient 21 6.5 8.0 Patient 22 5.5 7.1 Patient 23
5.8 7.4 Patient 24 7.8 9.6 Patient 25 8.3 10.1 Patient 26 7.8 9.8
Patient 27 6.1 8.2 Patient 28 8.3 10.4 Patient 29 7.4 9.7 Patient
30 8.3 10.9 Rate of patients with reduced blood phosphorus level:
13% (4 patients) Average increase in blood phosphorus level for
patients 1 to 30: 0.94 mg/dL Average increase in blood phosphorus
level for control group: 1.36 mg/dL
[0110] As shown in Table 7, it can be understood that 4 (patients 1
to 4) of the 30 patients had lowered blood phosphorus levels after
the ingestion of the killed bifidobacterium-containing seamless
capsule. On the other hand, lowering of the blood phosphorus level
was not observed after the ingestion of the killed
bifidobacterium-containing seamless capsule for the remaining 26
patients. The average increase in blood phosphorus level was 0.94
mg/dL for the 30 patients compared to the increment for the control
group of 1.36 mg/dL, and it can be understood that a blood
phosphorus level elevation was inhibited.
Example 6
[0111] 30 dialysis patients ingested the cell powder of
bifidobacterium (Bifidobacterium longum subspecies longum JCM1217)
as used in Example 1 without encapsulation in 2 billion cells of
bifidobacterium daily over 4 weeks. The condition of ingestion was
the same as that in Examples 1. The patients did not take any other
blood phosphorus level lowering agents. While under dietary
instructions by a dietitian, not all patients had the same meals.
Blood was collected before and 4 weeks after the beginning of the
ingestion of the cell powder of bifidobacterium, and blood
phosphorus levels were measured. Table 8 shows the results.
[0112] For 15 patients who did not take either the cell powder of
bifidobacterium or any other blood phosphorus level lowering agents
(control group), the average blood phosphorus level before
treatment was about 6.75 mg/dL, and the average blood phosphorus
level four-week after treatment was about 7.97 mg/dL (for the
control group, the average increase in blood phosphorus level was
1.21 mg/dL).
TABLE-US-00008 TABLE 8 Blood phosphorus level (mg/dL) 4 weeks after
Before the beginnig ingestion of ingestion Patient 1 6.3 6.1
Patient 2 6.3 6.2 Patient 3 5.9 5.9 Patient 4 8.0 8.0 Patient 5 6.3
6.4 Patient 6 5.7 5.9 Patient 7 7.8 8.2 Patient 8 6.3 6.7 Patient 9
7.2 7.7 Patient 10 6.6 7.2 Patient 11 7.7 8.4 Patient 12 6.3 7.0
Patient 13 8.5 9.2 Patient 14 8.3 9.2 Patient 15 7.0 8.3 Patient 16
7.8 9.1 Patient 17 5.9 7.3 Patient 18 7.2 8.6 Patient 19 6.3 7.7
Patient 20 8.2 9.7 Patient 21 7.7 9.3 Patient 22 6.5 8.3 Patient 23
8.1 10.1 Patient 24 8.5 10.6 Patient 25 7.4 9.6 Patient 26 6.8 9.0
Patient 27 5.6 7.9 Patient 28 8.1 10.4 Patient 29 6.4 8.7 Patient
30 5.5 8.1 Rate of patients with reduced blood phosphorus level: 7%
(2 patients) Average increase in blood phosphorus level for
patients 1 to 30: 1.15 mg/dL Average increase in blood phosphorus
level for control group: 1.21 mg/dL
[0113] As shown in Table 8, it can be understood that 2 (patients 1
and 2) of the 30 patients had lowered blood phosphorus levels after
the ingestion of the cell powder of bifidobacterium. On the other
hand, lowering of the blood phosphorus level was not observed after
the ingestion of the cell powder of bifidobacterium for the
remaining 28 patients. The average increase in blood phosphorus
level was 1.15 mg/dL for the patients compared to the increment for
the control group of 1.21 mg/dL, and it can be understood that a
blood phosphorus level elevation was inhibited slightly.
Example 7
[0114] In order to prepare the Bifidobacterium-containing seamless
capsule, the procedure was followed as in Example 1 except that the
cell powder of bifidobacterium (Bifidobacterium bifidum JCM1255)
was used in place of that of bifidobacterium (Bifidobacterium
longum subspecies longum JCM1217).
[0115] 20 dialysis patients ingested 0.2 g of the
bifidobacterium-containing seamless capsule thus prepared daily
under the same conditions of ingestion as in Example 1. Blood was
collected before and 4 weeks after the beginning of the ingestion
of the bifidobacterium-containing seamless capsule, and blood
phosphorus levels were measured. Table 9 shows the results.
[0116] For 15 patients who did not take either the cell powder of
bifidobacterium or any other blood phosphorus level lowering agents
(control group), the average blood phosphorus level before
treatment was about 6.75 mg/dL, and the average blood phosphorus
level four-week after treatment was about 7.97 mg/dL (for the
control group, the average increase in blood phosphorus level was
1.21 mg/dL).
TABLE-US-00009 TABLE 9 Blood phosphorus level (mg/dL) 4 weeks after
Before the beginnig ingestion of ingestion Patient 1 5.93 3.87
Patient 2 6.47 5.10 Patient 3 7.04 6.25 Patient 4 6.36 5.62 Patient
5 6.61 6.06 Patient 6 5.91 5.40 Patient 7 8.39 7.97 Patient 8 7.13
6.75 Patient 9 6.31 5.96 Patient 10 6.65 6.47 Patient 11 7.21 7.23
Patient 12 7.06 7.25 Patient 13 6.28 6.49 Patient 14 6.44 6.69
Patient 15 7.03 7.36 Patient 16 6.56 6.92 Patient 17 5.48 6.11
Patient 18 6.38 7.03 Patient 19 8.21 9.04 Patient 20 5.10 6.46 Rate
of patients with reduced blood phosphorus level: 50% (10 patients)
Average increase in blood phosphorus level for patients 1 to 20:
0.13 mg/dL Average increase in blood phosphorus level for control
group: 1.21 mg/dL
[0117] As shown in Table 9, it can be understood that 10 (patients
1 to 10) of the 20 patients had lowered blood phosphorus levels
after the ingestion of the bifidobacterium-containing seamless
capsule. On the other hand, lowering of the blood phosphorus level
was not observed after the ingestion of the
bifidobacterium-containing soft capsule for the remaining 10
patients. The average increase in blood phosphorus level was 0.13
mg/dL for the 20 patients compared to the increment for the control
group of 1.21 mg/dL, and it can be understood that a blood
phosphorus level elevation was inhibited.
[0118] For the control group, the blood phosphorus levels after
treatment were significantly higher than the levels before
treatment (Paired T-test: p<0.01), although no significant
increase was observed in the treated group. Therefore, it can be
understood that the bifidobacterium-containing seamless capsule
obtained in Example 7 significantly inhibit a blood phosphorus
level elevation.
Example 8
[0119] In order to prepare the seamless capsule containing lactic
acid bacterium, the procedure was followed as in Example 1 except
that the cell powder of lactic acid bacterium (Lactobacillus
acidophilus JCM1132) was used in place of that of bifidobacterium
(Bifidobacterium longum subspecies longum JCM1217).
[0120] 20 dialysis patients ingested 0.2 g of the lactic acid
bacterium-containing seamless capsule thus prepared daily under the
same conditions of ingestion as in Example 1. Blood was collected
before and 4 weeks after the beginning of the ingestion of the
lactic acid bacterium-containing seamless capsule, and blood
phosphorus levels were measured. Table 10 shows the results.
[0121] For 15 patients who did not take either the cell powder of
lactic acid bacterium or any other blood phosphorus level lowering
agents (control group), the average blood phosphorus level before
treatment was about 6.75 mg/dL, and the average blood phosphorus
level four-week after treatment was about 7.97 mg/dL (for the
control group, the average increase in blood phosphorus level was
1.21 mg/dL).
TABLE-US-00010 TABLE 10 Blood phosphorus level (mg/dL) 4 weeks
after Before the beginnig ingestion of ingestion Patient 1 6.17
4.48 Patient 2 6.61 5.21 Patient 3 5.74 4.50 Patient 4 8.79 7.63
Patient 5 7.13 6.45 Patient 6 6.55 5.90 Patient 7 6.43 6.00 Patient
8 7.24 7.04 Patient 9 8.92 8.76 Patient 10 6.44 6.47 Patient 11
7.07 7.23 Patient 12 6.20 6.43 Patient 13 5.76 6.05 Patient 14 6.77
7.29 Patient 15 5.69 6.30 Patient 16 7.12 7.74 Patient 17 6.29 6.95
Patient 18 6.30 7.00 Patient 19 6.15 7.56 Patient 20 5.18 6.73 Rate
of patients with reduced blood phosphorus level: 45% (9 patients)
Average increase in blood phosphorus level for patients 1 to 20:
0.04 mg/dL Average increase in blood phosphorus level for control
group: 1.21 mg/dL
[0122] As shown in Table 10, it can be understood that 9 (patients
1 to 9) of the 20 patients had lowered blood phosphorus levels
after the ingestion of the lactic acid bacterium-containing
seamless capsule. On the other hand, lowering of the blood
phosphorus level was not observed after the ingestion of the lactic
acid bacterium-containing seamless capsule for the remaining 11
patients. The average increase in blood phosphorus level was 0.04
mg/dL for the 20 patients compared to the increment for the control
group of 1.21 mg/dL, and it can be understood that a blood
phosphorus level elevation was inhibited.
[0123] For the control group, the blood phosphorus levels after
treatment were significantly higher than the levels before
treatment (Paired T-test: p<0.01), although no significant
increase was observed in the treated group. Therefore, it can be
understood that the lactic acid bacterium-containing seamless
capsule obtained in Example 8 significantly inhibit a blood
phosphorus level elevation.
Examples 9 to 17
[0124] Except that the cell powders of bifidobacterium
(Bifidobacterium bifidum JCM1255: Example 9), bifidobacterium
(Bifidobacterium animalis JCM10602: Example 10), bifidobacterium
(Bifidobacterium infantis JCM7007: Example 11), bifidobacterium
(Bifidobacterium dentium JCM1195: Example 12), lactic acid
bacterium (Lactobacillus casei JCM1134: Example 13), lactic acid
bacterium (Lactobacillus plantarum JCM11125: Example 14), lactic
acid bacterium (Lactococcus lactis subspecies lactis JCM7638:
Example 15), lactic acid bacterium (Enterococcus faecium JCM5804:
Example 16), and lactic acid bacterium (Enterococcus faecalis
JCM5803: Example 17) were used in place of that of bifidobacterium
(Bifidobacterium longum subspecies longum JCM1217), the procedure
was followed as in Example 1 in order to prepare the seamless
capsule containing Bifidobacterium or lactic acid bacterium.
[0125] 20 dialysis patients ingested 0.2 g of the seamless capsule
containing bifidobacterium or lactic acid bacterium thus prepared
daily under the same conditions of ingestion as in Example 1. Blood
was collected before and 4 weeks after the beginning of the
ingestion of the seamless capsule containing bifidobacterium or
lactic acid bacterium, and blood phosphorus levels were measured.
Table 11 shows the results.
TABLE-US-00011 TABLE 11 Rate of the patients with Average increase
reduced blood in blood phosphorus level phosphorus level Name of
strain used (%) (mg/dL) Example 9 Bifidobacterium bifidum JCM1255
55 -0.02 Example 10 Bifidobacterium animalis JCM10602 50 -0.10
Example 11 Bifidobacterium infantis JCM7007 40 0.01 Example 12
Bifidobacterium dentium JCM1195 40 0.19 Example 13 Lactobacillus
casei JCM1134 35 0.62 Example 14 Lactobacillus plantarum JCM11125
35 0.45 Example 15 Lactococcus lactis subspecies lactis 35 0.14
JCM7638 Example 16 Enterococcus faecium JCM5804 45 0.21 Example 17
Enterococcus faecalis JCM5803 30 0.26 Average increase in blood
phosphorus level for control group: 1.32 mg/dL
[0126] For 15 patients who did not take either bifidobacterium or
lactic acid bacterium or any other blood phosphorus level lowering
agents (control group), the average blood phosphorus level before
treatment was about 6.48 mg/dL, and the average blood phosphorus
level four-week after treatment was about 7.59 mg/dL (for the
control group, the average increase in blood phosphorus level was
1.32 mg/dL).
[0127] As shown in Table 11, it can be understood that Examples 9
and 10 had lowered blood phosphorus levels after the ingestion of
the seamless capsule containing bifidobacterium. On the other hand,
lowering of the blood phosphorus level was not observed after the
ingestion of the seamless capsule containing bifidobacterium or
lactic acid bacterium for Examples 11 to 17. Compared to the
increment for the control group of 1.32 mg/dL, it can be understood
that a blood phosphorus level elevation was inhibited.
[0128] For the control group, the blood phosphorus levels after
treatment were significantly higher than the levels before
treatment (Paired T-test: p<0.01), although no significant
increase was observed in the treated group. Therefore, it can be
understood that the bacterium- or the lactic acid
bacterium-containing seamless capsule obtained in Examples 9 to 16
significantly inhibit a blood phosphorus level elevation.
[0129] It is clear from the experimental results presented above
that the lactic acid bacteria have the effect of inhibit a blood
phosphorus level elevation. Furthermore, it can be understood that
the lactic acid bacteria, when ingested in the form of an enteric
capsule, remarkably inhibit a blood phosphorus level elevation.
INDUSTRIAL APPLICABILITY
[0130] The inhibitor for blood phosphorus level elevation of the
present invention is highly safe, is readily administerable, and
can sufficiently inhibit a blood phosphorus level elevation.
Therefore, the inhibitor for blood phosphorus level elevation of
the present invention is applicable to a food product, a
nutritional supplement, or the like that can inhibit a blood
phosphorus level elevation for people such as dialysis patients and
patients with impaired renal function.
* * * * *