New Material Proposal

Fucoidan from Okinawa Mozuku

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Kanehide Bio Co., Ltd

Presentation 2／22

What is Okinawa Mozuku?
Okinawa Mozuku Cladosiphon okamuranus
Mozuku is a brown alga which is classified under the family Chordariaceae and is an original
species in only Okinawa islands. Eating mozuku habit has a long history however it was only
for special areas and a period until established a farming mozuku skill even if in Okinawa. The
farming skill had been developed after 1970’s and now farmed 20 thousands tons of mozuku in
Okinawa in a year, this amount means 90% in gross domestic production. Comparatively
mozuku and other seaweeds, mozuku has a higher content of Fucoidan and further lower
content of impurity as like alginic acid. This is to say Okinawa mozuku is excellent as amaterial of Fucoidan.
2.5

Harvesting Okinawa Mozuku

Content of Fucoidan in each seaweeds

2

By wet algae

1.5

沖縄産
98.8 %

1

0.5

0

A nursery of Mozuku

Okinawa
MozukuIto
MozukuEdible
brown
algae

kelp

WakameRefference ：でん粉と食品 第29号「長寿と海藻」

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Kanehide Bio Co., Ltd

Presentation 3／22

Seaweed-eating culture in Okinawa
For Okinawan people which are surrounded by sea, seaweed is a important food source.
Particularly in the spring tide we often gather seaweeds and eat them, we call this culture“hamakudari” in Japanese. Not only Okinawa, in Japanese meals, more than 20 species areused including a seaweed-based soup stock however these seaweeds are not eaten. In
Okinawa we eat seaweeds with flied or pickled consequently we can take all of usefulcomponent of seaweeds. This may be a cause of the longevity island.

Okinawan people gathering seaweeds.

Okinawan traditional foods.

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Kanehide Bio Co., Ltd

Presentation 4／22

What is Fucoidan?
Fucoidan is a soluble viscosity polysaccharide
which is contained in brown algae as like Mozuku,
sea tangle and wakame. Compared with otheralgae, Mozuku has higher content and higher
purity of Fucoidan.

History of Fucoidan studies
1913
After 1950’s
After 1960’s

After 1970’s
1996

Sulfate group

FucoseGlucuronic acid

Constitutional formula of Fucoidan extract from Mozuku：Dr. Kylin from Uppsala University discovered Fucoidan into brown algae.
：Reported in blood anticoagulant activities of Fucoidan.
：Studied in component of Fucoidan from variety brown algae and chemical
constitution.
：Reported in anti tumor activities.
：Dr. Tako from Ryukyu University reported that Okinawa mozuku contain a lot of
Fucoidan.

Bioactivity of Fucoidan based on studies
1. Anti tumor activity
2. Apoptosis induction activity against cancer cell
3. Immunostimulatory activity
4. Hypoglycemic activity
5. Blood pressure lowering effect6. Anti-inflammation activity(for atopic dermatitis)
7. Antiviral activity (for WEE and HSV)All rights reserved.

Kanehide Bio Co., Ltd

Kanehide Bio Fours points of Fucoidan
1．Using only Mozuku from Okinawa.
2．Fucoidan is high purity and produced with our original manufacturing method.
3．Nothing used any dextrin, 100% pure powder(we guarantee Fucoidan and sulfate groupcontent)
4．Substantial evidence and guarantee of safety by human trials
5．Manufacturing products in a factory certified ISO22000 and GMP.
6．We have a lot of experience to export and domestic sell.

Manufacturing process

Receipt of materialContinuous production in the Kanehide plant.

Extraction by organic acid
Separation of solid and liquid
Ultrafiltration(salt and iodine)
Concentration

First inspection
Powderization
Second inspection( dosage form: powder, bulk)

Pockaging ( dosage form: tablet, granule, cuspule, liquid )Final Inspection
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Kanehide Bio Co., Ltd

Presentation 6／22

Chemical analysis of Fucoidan from Mozuku
Item

Fucosecontent 1)

Uronic acidcontent 2)

Sulfate groupcontent 3)

Molecularmass 4)

Fucoidancontent 5)

Measuredvalues6)

45.6 %

26.7 %

19.1 %

77,00089.9 %

Value ofstandard

-

-

≧13 %

30,000～150,000>85 %1) Anthrone-sulfuric acid method 2）Carbazole sulfuric acid method
3) Ion chromatography4) HPLC gel filtration technique5）HPLC gel filtration technique 6）Average of 4 lots
Species of seaweeds

Characteristics of ingredient

Okinawa mozuku

Higher content of fucose compared with other seaweeds.
It occupy 40% of sulfate group in fucose.

Kelp

More than 3 kinds of polysaccharide as Fucoidan.
It contains more than 25% of sulfate group.

WakameIt contains as amount as fucose and galactose which contains Sulfatedfucogalactan.

MekabuIt contains Sulfated polysaccharides which is a similarity with Wakame.
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Kanehide Bio Co., Ltdプレゼンテーション資料 7／22

Bioactivation of Fucoidan from Okinawa Mozuku
1. Apoptosis inducing activity
2. Killing tumor cells activity
3. Anti-tumor activity (animal test)

4. Macrophage activation (immunostimulation)
5. Anti- atopic dermatitis activity(animal test)
6. Anti-obesity activity(animal test)
7. Immunostimulation activity (clinical examination)

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Kanehide Bio Co., Ltdプレゼンテーション資料 8／22

Apoptosis Inducing activity【Test Method】
Added Fucoidan to HL60（ promyelocytic leukemia cell strain) and NOS4(cultured human ovarian cancer cellstrain), evaluated the ratio of DNA fragmentation in cells as apoptosis inducing potency.
40

40
Mozuku fucoidan

Apoptosis Inducing potency(% DNA Fragmentation)

Apoptosis Inducing potency(% DNA Fragmentation)

35

Kelp fucoidan

30
25

20
15

10
5

Mozuku fucoidan

35

Kelp fucoidan

30
25
20
15

10
5
0

0
10

100

1000

Concentration（μg／mL)
Apoptosis induction of promyelocytic leukemia cellstrain（HL60)

10

100

1000

Concentration （μg／mL)

apoptosis inducing of cultured humanovarian cancer cell strain（NOS4)【Result】
Measuring the apoptosis inducing activity with Fucoidan from Okinawa Mozuku and from kelp, it was found that
Fucoidan from Okinawa Mozuku had a more significant apoptosis inducing effect than Fucoidan from kelp.
The data from result of joint research with Tokyo University of Science

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Kanehide Bio Co., Ltd

Presentation 9／22

Killing tumor cells activity【Test Method】
Using PBL (human lymphocyte) as normal cells and Molt-4( acute lymphoblastic leukemia) as
continuous cancer cell from human, measuring a survival rate after adding Fucoidan.
160

The survival rate of normal cells
(%)

140
120
100

Molt4
PBL

80
60
40

Added
FucoidanNo added
Fucoidan

20
0

0

500
1000
1500
Concentration
添加濃度（μg/mL）

2000

GRAPH. The effect of Fucoidan with PBL and Molt-4pic.) The comparison of multiplication of Molt-4 withno added fucoidan and added fucoidan【Result】
It is not found a viability loss of normal cells (PBL) whereas the survival rate of cancer cells ( Molt4 ) was controlled depending on concentration. As a result we suppose Okinawa mozuku fucoidan tohave a specific effect against cancer cells.
The data by Kanehide Bio Co., Ltd.
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Kanehide Bio Co., Ltd

Presentation 10／22

Anti-tumor activity (animal test)【Test Method】
Tested the tumor cells (mouse sarcoma cell strain :Sarcoma180) growth inhibition with nude mouse. afteradministering cancer cells in subcutaneous. Oral inoculating 100 mg per kg of Okinawa mozuku fucoidan for 21
days once per day after administering cancer cells in subcutaneous.
n=
n=
4000
7.00
3
3
5.59
6.00

モズク由来フコイダン
Okinawa
mozuku fucoidan

5.00

weightCancer腫瘍重量（ｇ）

3
volumeCancer腫瘍容積（mm
）

3000

対照群
Control

2000

1000

4.00
3.00

1.94

2.00
1.00

0
0

7

14

Effect against cancer growth

21 days
期間

0.00

control
対照群

Okinawa
mozuku fucoidan
モズク由来フコイダン

Effect against cancer weight （21days）【Result】
It is verified a significant difference （p＜0.005）of Okinawa mozuku fucoidan compared with the control on the 14th day and on
the 21st day verified a significant difference （ｐ＜0.0005）.as a result of the test, it is verified that Okinawa mozuku fucoidan
controls a growth of cancer cells.
The data from result of joint research with Prefectural University of Kumamoto

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Kanehide Bio Co., Ltd

Presentation 11／22

Macrophage activating potency (immunostimulation)【Test Method】
Macrophage is a cell which has a excellent phagocytic activity, it plays an important part likewise not only digesting
foreign matters in a body, but also releasing cytokines or active oxygen, acting to bridge and accommodating toimmunoreaction such as antigen presentation, attacking or removing cancer cells and so on.
In addition it is known as glucose metabolism system participate when macrophages produce active oxygen.
Consequently we can estimate that the higher consumption of glucose, the higher production quantity which means thehigher immunostimulating activity.
）
Consumption of glucose (mg/well140

Added 100μg/mL of fucoidan

120
100
80
60
40

Before20001020 30 40 50 60 70 80 90 100
Sample concentration（μg／mL）

After

It is observed spreading dilatation which we
can see when macrophage activates.

Data: Activation of macrophage
indicated by consumption of glucose【Result】
Measuring activation of macrophage indicated by consumption of glucose, we confirmed that
Okinawa mozuku fucoidan has a enhancing glucose consumptive activity which means Okinawa
mozuku fucoidan has immunostimulating activity.
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Kanehide Bio Co., Ltd

Presentation 12／22

Anti atopic dermatitis activity (animal test)【Method】

NC/NgaTnd Crj
4W old ♂

Inspection Item
1. Pruritus cutaneous
2. Redness and bleeding
3. Edemas
4. Excoriation and tissue deficit
5. Incrustation and dryness

・・・・・
inducing inducing

inducing

sensitization

Feeding Fucoidan（14days）

Additional inducing（after determination）

Determination
Categorizing 4 steps from asymptomatic to high symptom and determining chronologically the average value of total
rating as seriousness.
7
Control
Fucoidan＊

6

Average value

5
評
点 4
平
均 3
値
2

A

B

1
0
pre.

3

7

10

観察時間（日）Days observed

14Pic) influences regard to atopic dermatitis
A: Starting day
B: Day 14Pic) influences regard to atopic dermatitis.
＊：Compared with control, there was p＜0.05 of significant difference.

【Result】
Determined improvement for dermatitis with oral administration of fucoidan in atopic dermatitis mice.
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Kanehide Bio Co., Ltd

Presentation 13／22

Anti-obesity activity (animal test)
350

50
0

1.0

2.0

3.0

4.0

5.0

0

60

120

Concentration（mg/ml）

Inhibition of pancreatic lipase by
Fucoidan

180

240

300

360

Time（ｍｉｎ）

Suppression of neutral fat increases by
Fucoidan n=4

Preventing weight gain
【Result】
Maintained the weight of mouse andParametrial Adipose Tissue by adding
Fucoidan to high fat food.

Mouse's weight (g)
体重（ｇ）

45

36

27
Normal Food
普通食
High Fat Food
高脂肪食
High Fat Food+Fucoidan 3%
高脂肪食＋フコイダン3%High Fat Food+Fucoidan 5%
高脂肪食＋フコイダン5%High Fat Food+Fucoidan 10%18

高脂肪食＋フコイダン10%

2.50
1.81

2.00
1.50

1.00

0.98

0.94

1.00
0.50
0.00

9
0

1

2

3

4

5

6

7

8

9 10

Breeding period(week)

飼育期間（週）(pic.)Effect of Fucoidan on body weightin mouse with high fat food

図．高脂肪食マウスの
体重推移

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High fat food 3% Fucoidan 5% Fucoidan 10% Fucoidan

10
%フ
コ
イ
ダ
ン

50
0.0

100

フ
コ
イ
ダ
ン

60

Fucoidan extracted from Okinawa Mozuku inhibitedthe activity of pancreatic lipase which is a lypolytic
enzyme. In experiments of oral administration inrats, inhibited for rise in blood neutral fat afteradministration of lipid.

150(pic.)Effect of Fucoidan on parametrial
adipose tissue weight in mouse with highfat food

5%

70

200

フ
コ
イ
ダ
ン

80

Emulsification of a lipid＋Fucoidan

250

3%

90

300

Inhibition of pancreatic lipase activity andActivity of reducing neutral fat

高
脂
肪
食

100

Emulsification of a lipid

Parametrial組織重量（g）
adipose tissue weight (g)Blood plasme neutralfat（mg/dl）

Pancreatic lipase activity（％）

110

図．マウス子宮傍脂肪組織
重量に及ぼす影響

Kanehide Bio Co., Ltd

Presentation 14／22

Immunostimulation activity (human test : mucous immune in oral)
Secretory IgA （s-IgA）✓ Main immunoglobulin found in mucous secretions and it removes foreign
matters.
✓ The first line of defense against bacteria, food residues, yeast, parasites andviruses and it prevents invasion of them.
✓ In the oral, s-IgA in saliva defends against bacteria and viruses infect toupper airway.
Measuring amount and speed of s-IgA secretions as an index of oral
immune✓ Sampling saliva is simple, easy noninvasive.
✓ The measuring is a useful as a tool for observing condition of healthy
individual’s immune system.

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Kanehide Bio Co., Ltd

Presentation 15／22

Immunostimulation activity (human test : mucous immune in oral)
Research subjects✓ 22 healthy people（ 11 male, 11 female）✓ From 20 years old to 65 years old（average 42.3±10.4 years old）
✓ Slowing secretions of secretory IgA(s-IgA)

Test sample✓ Fucoidan in capsule✓ 5 capsules /day（contained 1,175mg of Okinawa mozuku extract)

How to test
✓ Taking 5 capsules every day for 12 weeks.
✓ Measuring speed of s-IgA secretions in saliva at the 4th week, the
8th week and the 12th week.
✓ Urinalysis and blood inspection on the 12th week.
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Kanehide Bio Co., Ltd

Presentation 16／22

Immunostimulation activity (human test : mucous immune in oral)
唾液中s-IgA分泌速度（μg/min）of IgA secretions in saliva (㎍/min)
Speed

400

350

300

＊＊

250

199

＊＊

＊＊

212

219

the ８週目
8th week

the１２週目
12th week

200

150

128

100

50

0

事前test
Beforethe４週目
4th week**p<0.01✓ Comparing with before test, the speed of s-IgA secretions in saliva at the 4thweek, the 8th week and the 12th week significantly increased.
✓ Expected immune boosting effect by taking fucoidan from Okinawa mozuku.
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Kanehide Bio Co., Ltd

Presentation 17／22

Safety (Data of clinical examination in before and after taking )＜Hematology test＞＜Biochemical test＞

Item

Unit ofmeasureStandard

Before

12th
week

WBC

(/μL)

3300 - 9000

6013.6 ±2065.95904.5 ±1839.9RBC(×104/μL)

Male：430 - 570
Female：380 – 500

471.3 ±
49.1

464.4 ±
45.9

Hb

(g/dL)

Male：13.5 – 17.5
Female：11.5 – 15.0

14.06 ±
1.69

13.87 ±
1.85(%)

Male：39.7 – 52.4
Female：34.8 – 45.0

43.49 ±
4.42

42.44 ±
4.60**

Ht

It is founded some data fluctuation andadverse event (as like cavities) however
the symptoms are mild and be detectedthat the symptoms don’t have relevanceto Fucoidan.

Item

Unit ofmeasure

AST
ALT
LDH

(U/L)
(U/L)
(U/L)

γGT

(U/L)

ALP
TB
TP
Alb

(U/L)
(mg/dL)(g/dL)(g/dL)

Cr(mg/dL)UN(mg/dL)UA(mg/dL)

GLU
TG
TC(mg/dL)(mg/dL)(mg/dL)

HDL-C(mg/dL)

Standard

Before12th week

10 - 40
5 - 45120 - 240
Male：80以下
Female：30以下
100 - 325
0.2 - 1.2
6.7 - 8.3
3.8 - 5.2
Male：0.61 - 1.04
Female：0.47 - 0.79
8.0 - 20.0
Male：3.8 - 7.0
Female：2.5 - 7.0
70 – 109
30 – 149
120 – 219
Male：40 - 85
Female：40 – 95
65 - 13919.9 ± 4.6
18.0 ± 7.2
166.9 ± 29.7

18.7 ± 5.0
16.3 ± 8.5
158.0 ± 28.5**

25.0 ± 13.3

22.4 ± 10.8*

213.4 ± 75.0
0.73 ± 0.23
7.39 ± 0.47
4.59 ± 0.29

209.4 ± 73.0
0.78 ± 0.29
7.28 ± 0.46
4.49 ± 0.31

0.758 ± 0.190

0.807 ± 0.180**

12.35 ± 2.90

12.52 ± 3.07

5.07 ± 1.18

5.21 ± 1.47

83.0 ± 5.6
83.3 ± 30.3
202.7 ± 40.8

84.0 ± 8.8
100.8 ± 76.7
188.0 ± 32.9**

65.4 ± 16.0

62.0 ± 15.9*(mg/dL)119.0 ± 32.2
107.0 ± 30.5*
LDL-C
the mean value ± standard deviation,**p<0.01, *p<0.05 vs before

Safety of Fucoidan from Okinawa Mozuku
22 healthy people approx. 1.2g/day×12 weeks・・・・NOTHING PROBLEM
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Kanehide Bio Co., Ltd

Presentation 18／22

Absorption of Fucoidan
Activity of Fucoidan
・Immunostimulation activity ・Anti tumor activity
・Anti virus activity ・Anti allergy activity
・Anti blood coagulating activity
・hypocholesterolemic activity and etc.,

Sulfategroup

Uronic acid
Fucose(pic.) Constitutional formula of Fucoidan from
Okinawa mozuku

Since Most of these reports are tests in vitro, absorbable is precondition for these
activity.

As Fucoidan is a macromolecule substance, generally considered most of Fucoidan
might not absorb intestinal.

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Kanehide Bio Co., Ltd

Presentation 19／22

Predicting absorbability in model testwith the small intestine membrane
Adding 1% of Fucoidan
Enteric side
（Apical）
Enteric side（Apical）
Basal membrane side（Basal）

Scheme of entericBasal membraneside（Basal）

Hydrolyzing
penetrated fucose
and analyzing by
HPLC method

Scheme of Caco-2 test system

Scheme of the test
Fucoidan
フコイダン

Fucoidan
フコイダン

0.00
0.50
1.00
1.50
2.00
2.50
Penetration
amount(Fucose
of
basal
side,
μg/cm2/h)
フコイダン透過量（Basal側フコース,μg/cm2/h）

フコイダンの透過量
Pic.) 図Penetration
amount for Fucoidan

0.000

0.020
0.040
0.060
0.080
Penetration
rate
after
2
hours(%)
2時間後の透過率（％）

0.100

図Penetration
フコイダンの透過率
Pic.)rate for Fucoidan【Result】
By Prospective study for absorbability of Fucoidan with intestine membrane model, we founded
Fucoidan through the small intestine membrane. It means we suggested Fucoidan is absorbablein intestinal .
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Kanehide Bio Co., Ltd

Presentation 20／22

Safety <Animal test and clinical examination ( Excess intake)>
4 week repeat dose studies using rats
Te s t f a c i l i t y：
Subjects：
Te s t S a m p l e ：Te s t m e t h o d ：CRO Rabiton Co., Ltd.
Wister rats, 10 each group( 5 male, 5 female )Fucoidan Extract Powder Capsule
1. 135mg/kg（low dosage）
2. 270mg/kg（high dosage）
Oral administration once in a day for 4
weeks✓ No abnormalities found in biochemical
examination of the blood orhematological assessment.
✓ No difference between groups
administered low dosage and highdosage.

Prolonged consumption tests of fucoidan
by healthy individuals
Te s t f a c i l i t y：
Subjects：

Te s t S a mp l e ：Tokyo Medical and Dental Univ.
Staff of Kanehide Bio Co., Ltd. andTokyo Medical and Dental University, 35
healthy people (19 male, 16 female)
Fucoidan Extract 50mL/bag (6g of
Fucoidan)1 bag per day for 1 to 3 month

Te s t m e t h o d ：✓ There was no significant difference in bodyweight, blood tests, or urinalysis tests before andafter taking Fucoidan. No adverse experienceswere recorded.
✓ Immune function tended to be elevated.

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Presentation 21／22

Confirmed evidence (conference presentation)
Year

Title of the presentation and name of the event
・Japanese Cancer Association 62nd General Meeting“Potential T cell Leukemia Treatments with Fucoidan from Cladosiphon okamuranus ”

2003

・American Society of Hematology 45th General Meeting“Apoptosis Induced by Fucoidan from Cladosiphon Okamuranus Tokida in HTLV-Ⅰ-Infected T-Cell Lines and Primary
ATL Cells.”
・2nd Young Researchers Forum on Infectious Diseases“Potential T cell Leukemia Treatments with Fucoidan from Cladosiphon okamuranus ”

2004

・Japanese Cancer Association 63rd General Meeting“Anticancer effect of Fucoidan from Okinawan Cladosiphon okamuranus on Gastric Cancer Cell Lines”“Inverstigation of the Anticancer Effect due to differences in Molecular Weight and Elements of Fucoidan from
Okinawan Cladosiphon okamuranus ”・Japanese Cancer Association 64th General Meeting“Anticancer effect of Fucoidan from Okinawan Cladosiphon okamuranus on Sarcoma Cell Lines in Mice”

2005

・Japan Society Nutrition Food Science 59“A factor of Fucoidan from Okinawa mozuku relating to immunology”

2006

・Japanese Cancer Association 65nd General Meeting“Fucoidan extracted from Okinawan Cladosiphon okamuranus and the Mechanism for Inducing Mouse
Macrophage NO Production”“Anticancer effects of Okinawan Cladosiphon okamuranus”
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Presentation 22／22

Confirmed evidence (scientific reports and theses)
Year

Title of reports or theses

2004

・Vol. 66, No.1・2, The Japanese Journal of Constitutional Medicine“Anti-obesity Effects of Fucoidan Prepared from Cladosiphon Okamurananus tokida ( Okinawamozuku)

2005

・Nutrition and cancer, 52(2), 189-201「Fucoidan extracted from Cladosiphon okamuranus Tokida induces apoptosis of human T-cell leukemia virus type1infected T-cell lines and primary adult T-cell leukemia cells」

2005

・Japan Society Nutrition Food Science 58: 273-280(2005)“Biological Properties of Fucoidans Differing in Molecular Weight and Composition: Effects on Lymphocyte
Proliferation and Immunological Function”

2011

・Antiviral Therapy 2011;16:89-98「Fucoidan therapy decreases the proviral load in patients with human T-lymphotropic virus type-1-associatedneurological disease」

2012

・World J Gastroenterol 2012 May 14;18(18):2225-2230「Beneficial effects of fucoidan in patients with chronic hepatitis C virus infection」

2012

・INTERNATIONAL JOURNAL OF ONCOLOGY 40: 251-260, 2012
「Anti-tumor activity of fucoidan is mediated by nitric oxide released from macrophages」

2013

・Marine drugs, 11, 4267-4278; doi: 10.3390/md11114267「Cytotoxic Effects of Fucoidan Nanoparticles Against Osteosarcoma」

2018

・INTERNATIONAL JOURNAL OF ONCOLOGY 53: 801-814, 2018
「Mitotic kinase PBK/TOPK as a therapeutic target for adult T-cell leukemia/lymphoma」
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Kanehide Bio Co., Ltd