Enzymatic Hydrolyzed Brown Seaweeds...Enzymatic Hydrolyzed Brown Seaweeds 1. Sulfated Polysaccharides from Brown Seaweed 2. Small Molecular Fucoidan2014-10-05 Mr. Chia-Chin Sheu, President,

Enzymatic Hydrolyzed Brown Seaweeds

1. Sulfated Polysaccharides from Brown Seaweed2. Small Molecular Fucoidan

2014-10-05

Mr. Chia-Chin Sheu,President, Simpson Biotech Co., Ltd.

For Internal Evaluation Only, All Right Reserved

Simpson BiotechA Bio-Fermentation Company


An R&D oriented and innovative bio-fermentation company established in Dec. 1998 68 years of fermentation experiences (derived from Kong-Yen Vinegar Group)

Employees: 34 Capital: 10 millions US Dollars

Major Businesses Areas:Medicinal fungus/mushroom fermentation and their applications.Purification, identification, and the applications of the active ingredients isolated from the fermented medicinal fungus mycelia.Bio-transformation & Bio-conversion to produce functional ingredientsUse HERPEPSTM to produce recombinant proteins including industrial enzymes, pharmaceutical proteins and biosimilarsOEM/ODM services, i.e., producing tailor-made recombinant proteins/enzymes by using HERPEPSTM platform

About Simpson Biotech


SimpsonPure Bioactive Natural Ingredients Part 1: Fermented mycelia of Antrodia cinnamomea (A. camphorata)

Part 2: Fermented Cordyceps sinensis Mycelia & Extracts

Part 3. The Ginsenoside Compound K & the Fermented Ginseng

Part 4:

• Water Soluble/Low Molecular Chitosan (COS-LM) – Anti-microbial Agent

• Oligo Chitosan (COS-600) - the Chitobiose & Chitotriose

Part 5. Low-bitterness & Low Mw Peptides:

• Fish Scale Collagen Peptides

• Soy Peptides and Low Salt Seasoning Solutions

• Silk Peptides & others

Part 6. Enzymatic Hydrolyzed Seaweeds: Brown, Red & Green Algae

• Sulfated Polysaccharides from Brown Seaweed – Dietary Supplement & Drinks

• Small Molecular FucoidanFor Internal Evaluation Only, All Right Reserved

Part 6The Enzymatic Hydrolysation of

Seaweed into Low Molecular Functional Polysaccharides& Other Active Ingredients

- The Sea Weed Hydrolyzation Platform Technology


Seaweed polysaccharide hydrolyzation technology – for Brown, red & green seaweeds

Red Seaweed：

Brown Seaweed：

Green Seaweed：

www.topnews.in/usa/algae-knew-about-quantum-m...

www.liveaquaria.com/product/prod_display.cfm?..

oceanexplorer.noaa.gov/.../media/sargassum.html


What is Fucoidan?

1. Fucoidan is a sulfated polysaccharide (MW: average 20,000) found mainly in various species of brown seaweed such as mozuku, kombu, limu moui, bladderwrack, wakame, and hijiki

2. contains substantial percentages of L-fucose and sulfate ester groups, mainly derived from brown seaweed

3. Fucoidan is just a general term for the polysaccharide composed of fucose and sulfate ester plus galactose, annose, xylose, uronic acids, etc. 其化學成份為以硫酸岩藻糖為主的多醣體，結合岩藻糖以外的Galactose、Annose、Xylose、Uronic Acids等。同一構造的物質不能全稱做Fucoidan，它是岩藻糖鏈的總稱，至今還無法定出Fucoidan褐藻糖膠的定義。(Robertet al., 2004)。

Major interesting: antitumor and immunomodulatory


Structure of Fucoidan

褐藻醣膠 fucoidan oligosaccharide


資料來源：Mar. Drugs 2011, 9, 196-223

Fucoidan oligosaccharide


Biological activities of Sulfanted Polysacchrides from Brown Seaweed

Diabetes Control: α-Amylase & α-Glucosidase inhibitor for Type 2 diabetes. Anti-Anticoagulant and antithromboticAnti-virusAnti-tumor and immunomodulatoryAnti-inflammatoryBlood lipids reducingAntioxidant activitiy against hepatopathy, uropathy and renalpathyGastric protective effects


Previous Research on Seaweed & Seaweed Polysacchrides -1

Anticoagulant effects:Carbohydr. Res. 2007, 342, 2326-2330.Glycobiology 2005, 15, 369-381.

Anti-viral activity：Carbohydr. Res. 2007, 342, 2326-2330.Glycobiology 2005, 15, 369-381.

Anti- Tumor activity Am. J. Hematol. 2004, 78, 7-14.Bull. Exp. Biol. Med. 2007, 143, 730-732.Anticancer Res. 2005, 25, 2129-2133.

Immuno-regulation：J. Health Sci. 2005, 51, 394-397.Immunol. Lett. 2008, 115, 138-143.J. Med. Food 2005, 8, 446-453.


Anti-oxidation：Chin. Trad. Herbal Drugs 2003, 34, 824-826.J. Appl. Phycol. 2007, 19, 153-160.High Tech. Lett. 2005, 11, 91-94.Int. J. Biol. Macromol. 2008, 42, 127-132.

Hypolipidemic effect:Food Sci. 1999, 20, 45-46.Food Sci. 2001, 22, 92-95.J. Ocean Univ. Qingdao 2004, 34, 560-564.

Anti-inflammation activity：Int. Arch. Allergy Immunol. 2005, 137, 289-294.Biochem. Biophys. Res. 2006, 346, 345-350.

Liver-protective activity:Hepatol. Res. 2006, 35(3), 190-198.Pharmacol. 2008, 580, 380-384.

Previous Research on Seaweed & Seaweed Polysacchrides -2


Simpson Biotech’s Enzymatic Hydrolysation Process

Brown Seaweed Material wash Grinding add water & food grade enzymes mixing & reaction Sterilization reduce to room temperature press/filtration Hydrolzyted Clear Broth Pasteurization Frozen StorageBS Hydrolyzed Conc. BSH Conc.

Hydrolyzed Clear Broth Concentration add 4x Alcohol to get precipitation (BS Polysacchrides Powder, BSPS) & Clear broth without polysacchrides (Non-Poly Broth) Non-Poly Concentrated to get Non-Poly Conc. Liquid, BSNoPS Conc. Pasteurization Frozen Storage.

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Molecular Wt. Distribution (HPLC/GPC, RI detector) of Simpson Biotech Enzymatic Hydrolyzed Brown Seaweed (EHBS) and Commercial products善笙酵素水解褐藻多醣與市售產品之分子量比較圖

0.5Kd

善笙酵素水解褐藻多醣兼具有市售高分子多醣(日本T廠800~1200Kd)及小分子寡糖(台灣H廠0.5Kd)之特性，且獨具有特殊中分子多醣(10~50Kd) ，為其他廠牌商品所沒有。

Simpson EHBSJapan T BrandTaiwan H BrandAustralia M BrandJapan S Brand

0.5Kd

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Immunoregulatory Activities (Antibody IgM stimulation) of Polysaccharides from Simpson Biotech’s Enzymatic

Hydrolyzed Brown Seaweed (PS-EHBS)

Fig 1. PS-EHBS effectively induce the antibody (IgM) secretion in a human hybridoma cells HB4C5 assay

0

200

400

600

800

1000

1200

*

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Concentration (micro g/mL)

IgM

con

cent

ratio

n(ng

/ml)

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Immunoregulatory Activities (Anti-inflammation) of Polysaccharides from Simpson Biotech’s Enzymatic

Hydrolyzed Brown Seaweed (PS-EHBS)

ClaAPS

Contr

ol 0 125 250 5000

10000

20000

30000

40000

LPS (1 g/mL)

#

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Concentration (μ g/mL)

TNF-

α c

once

ntra

tion(

pg/m

l)

ClaAPS

Contr

ol 0 125 250 500

0

5000

10000

15000

20000

LPS (1 g/mL)

#

*****

Concentration (μ g/mL)

IL-6

con

cent

ratio

n(pg

/ml)

Fig 2. PS-EHBS effectively reduce the LPS-induced Superoxide in a murine RAW 264.7 macrophage cell line assay

Fig 3. PS-EHBS effectively inhibit both the cytokines TNF-α and IL-6 in a LPS-stimulated RAW264.7 macrophage cell line assay

0.0

0.2

0.4

0.6

LPS (1g/mL)

#

******

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**

Concentration (?g/mL)

Supe

roxi

de (O

D. 6

30 n

m)

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-

10.0

20.0

30.0

40.0

50.0

60.0

70.0

ControlPS-EHBS-A61.7mg/Kg

NK

cyt

olys

is, %

.

E/T=25E/T=50E/T=100

******

************

****

*** * *

PS-EHBS-A205.6mg/Kg

PS-EHBS-F205.6mg/Kg

EHBS-A205.6mg/Kg

Immunoregulatory activity (Natural Killer) of 2 different (A & F) Enzymatic Hydrolyzed Brown Seaweeds (EHBS)

and its Polysaccharides (PS-EHBS) on Miceat the Dosages Levels Equal to 0.3 and 1.0g/day/person

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0.0%

10.0%

20.0%

30.0%

40.0%

50.0%

60.0%

70.0%

80.0%

90.0%

100.0%

Control

Pha

gocy

tosi

s, %

.

MOI=0MOI=1MOI=5MOI=10

*

*

** ** **

Multiplicity of infection

PS-EHBS-A61.7mg/Kg

PS-EHBS-A205.6mg/Kg

PS-EHBS-F205.6mg/Kg

EHBS-A205.6mg/Kg

Immunoregulatory activity (Mononuclear Phagocytic) of 2 different (A & F) Enzymatic Hydrolyzed Brown Seaweeds

(EHBS) and its Polysaccharides (PS-EHBS) on Miceat the Dosages Levels Equal to 0.3 and 1.0g/day/person

α-Amylase & α-Glucosidase inhibition Activity of Simpson Biotech’s Enzymatic Hydrolyzed Brown

Seaweeds Polysaccharide & Commercial Products

Enzymes Inhibition Activities

α-Amylase Inhibition (IC50)

α-Glucosidase Inhibition (IC50)

Cebolixs by InSea2 52.8μg/ml 75.0μg/mlSimpson Hydrolyzed Brown Seaweed #1 27.4μg/ml 156μg/ml

Simpson Hydrolyzed Brown Seaweed #2 6.36μg/ml 83.9μg/ml

Commerical Fucoidan Ingrident #1 no activity

few activity (22% inhibition in 5mg/mL)

Commerical Fucoidan Ingrident #2 no activity no activity

Commerical Fucoidan Product #3 no activity no activity


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Starch Management: (α-Amylase inhibition)Clinical Trials for 23 healthy participants1. Reduce 48% blood glucose2. Reduce 12% insulin secretion3. Increase 8% Cederholm index (insulin sensitivity)

Competing Commercial Product - InSea2®- Active ingredients isolated from similar brown seaweeds

Sugar Management: (α-glucosidase inhibition)Clinical Trials for 42 healthy participants)1. Reduce 39% blood glucose2. Reduce 7% insulin secretion3. Increase 5% Cederholm index (insulin sensitivity)

Weight Management: Clinical Trials for 39 healthy (BMI 24~40) participants1. Increase 33% fullness2. Reduce 14% ingested calories3. Significant weight loss4. Safety: well tolerated for 3 month at daily

dosage = 1000 mg

The extraction yield (EY) & crude polysaccharides(CPP) molecular weight distribution (MWD) of all kind of

seaweeds (1.Green algae 海葡萄 )


The extraction yield (EY) & crude polysaccharides’ molecular weight distribution (MWD) of all kind of seaweeds

(2. Red algae 紫菜 )



(3. Brown algae 海带 )



(4. Brown algae 马尾藻 )



(5. Brown algae 裙带菜 )



(6. Brown algae 昆布 )


Thanks …Q&A ?

Contact to: Mr. Chia-Chin Sheu (許嘉欽)Simpson Biotech Co., Ltd. 善笙生物科技股份有限公司

[email protected]:+886-3-396-3466


Documents

Enzymatic Hydrolyzed Brown Seaweeds...Enzymatic Hydrolyzed Brown Seaweeds 1. Sulfated Polysaccharides from Brown Seaweed 2. Small Molecular Fucoidan2014-10-05 Mr. Chia-Chin Sheu, President,