1. Concept Cosmeceuticals
SP1- Growth Factor S P 1
: Skin
: permeable
: Want
Introduction of Cell Transducible Growth Factor for Skin Renewal SP1-EGF & SP1-FGF1
Real Effective Cosmeceuticals
begin at
Bioceltran

2. Development of Cell Transducible Growth Factors
Aim of the Study
Development of Cell Transducible Growth Factors
1) Skin Permeable Epidermal Growth Factor (SP1-EGF)
2) Skin Permeable Acidic Fibroblast Growth Factor(SP1-FGF1)
3) Skin Permeable Superoxide Dismutase(SP1-SOD)
4) DDS technology of skin permeable therapeutic humanized antibodies
Skin Care
Skin Regeneration
Treatment of Burn Injury
Before After
Before After
We aim to develop cosmeceuticals which cover from skin care, skin regeneration to burn injury through the development of cell transducible growth factors with PTD technology.

3. The Superiority and Creativity of PTD Technology
- Increase skin regeneration by improving skin permeability
No treated EGF
Product
Liposome linked EGF
PTD Conjugated EGF
Cosmetics containing even very small amounts of EGF, FGF1 is quite expensive in the current market. Notwithstanding the high price, protein itself is very difficult to permeate through the skin. So liposome formulated one is widely used. However liposomal formulation also has its limitation.
Thus we improved those limitation with PTD(Protein Transduction Domain) Technology.
As shown in the figure, PTD conjugated product (protein) showed better permeability compared to existing liposome. (Green fluorescence represent protein permeated deeply into the skin)
Comparison with the third party product

4. EGF (Epidermal Growth Factor)
1. Structural Characteristics
Consists of 53 amino acids
Molecular weight 6.2 kDa
3 S-S bond(Disulfide bond)
(6-20, 14-31, 33-42)
2. Function
Anti-aging & Improve Pre-made winkles by skin regeneration
Improve skin tone
Soft skin, Brightening and Rejuvenation
Remove skin scar by forming new epidermis
FGF1 (Acidic Fibroblast Growth Factor)
1. Structural Characteristics
Consists of 155 amino acids
Molecular weight 17.3 kDa
2. Function
Prevent winkles, aging, dry skin by synthesizing Collagen, Elastin and Glucosaminoglucan under the skin.
Promote skin elasticity by acting on the cells in the dermis.

5. Expression of SP1-EGF & SP1-FGF1
SDS-PAGE
Western blot
(A) SP1-EGF
(B) SP1-FGF1 M 2 1
SP1-EGF and SP1-FGF1 were analyzed by 15% SDS-PAGE and Western bloting using specific antibodies for EGF and FGF1.
M : Size marker
: Before the expression of SP1-EGF & SP1-FGF1
: After the expression of SP1-EGF & SP1-FGF1
# SP1-EGF molecular masses : 12kDa, # SP1-FGF1 molecular masses : 19kDa

6. Phospho EGF receptor (Tyr1068)
Activity of SP1-EGF & SP1-FGF1
(A) SP1-EGF
SP1-EGF 10
100ng/ml
Phospho EGF receptor (Tyr1068)
(B) SP1-FGF1
SP1-FGF1
Phospho FRS2-alpha
(Tyr196)
Used Cell Line A431
Used Cell Line NIH3T3
The data above are bioactivity test by specific growth factor marker of SP1-EGF and SP1-FGF1
The data above shows the dose dependent activity of SP1-EGF, SP1-FGF1 by showing phosphorlation status of EGF & FGF1 receptor as low as 10ng/ml concentration.

7. Effects of SP1-EGF & Sp1-FGF1 in cell proliferationB C D
Used Cell Line : NIH3T3
Treated Concentration : each 100ng/ml
A : Control
B : SP1-EGF
C : SP1-FGF1
D : SP1-EGF + SP1-FGF1 Mixture
The data above shows the proliferative activity of SP1-EGF, SP1-FGF1 in cells(NIH3T3). Compared to control A, all of the growth factors showed proliferative effects.

8. SP1-EGF + SP1- FGF1 Conc. (ng/ml)
Effects of SP1-EGF & Sp1-FGF1 in cell proliferation
SP1-EGF Proliferation
SP1-EGF Conc. (ng/ml)
SP1-EGF + SP1- FGF1 Conc. (ng/ml)
SP1-FGF1 Proliferation
SP1-FGF1 Conc. (ng/ml)
Combination
Bioceltran
The figure above is graph of the proliferative activity of SP1-EGF & SP1-FGF1 in cells(NIH3T3).
There is dose dependent proliferative effects from the concentration of 0.1ng/ml.

9. Effects of SP1-EGF & Sp1-FGF1 in Collagen Synthesis
Combination
Control
Ratio % vs GAPDH
Used Cell Line : CCD-986SK
Treated Concentration : each 100ng/ml
Combination : SP1-EGF + SP1-FGF1 Mixture
The graph above represent the ability of SP1-EGF and SP1-FGF1 in collagen synthesis. Collagen is distributed and acted in the dermis layer of the skin. SP1-FGF1 showed best collagen synthesis when measured its messenger RNA levels.

10. Degrees of Skin permeability
A : Control, B : Normal EGF, C : SP1-EGF, D : Normal FGF1, E : SP1-FGF1
Used skin : Porcine skin
Treated Concentration : each 100ng/ml
Treated Time : 3hr
Epidermis
The results above represent microscopically observed cross section of skin after fluorescent labeled SP1-EGF, SP1-FGF1 treatment to see the degree of tissue permeability .
In contrast to other EGF and FGF1, SP1-EGF and SP1-FGF1 showed superior permeability compared to others.

11. (INCI name : processing) Lyophilized or Solution stock
SP1-EGF
(Skin Permeable - Epidermal Growth Factor)
Characteristics of SP1-EGF
PTD & EGF Fusion Protein
The PTD fusion protein with EGF makes skin permeability better than with protein itself so maximize its effects by deep penetration into the cells.
High Purity and High Activity
With purity more than 95% , suitable as a cosmetic
Ingredient and more active than normal EGF.
Safety and Stability
Free from various microbial contamination and allergen. Suitable for the specification. Through the toxicity test, safety is assured. It is proved to be stable up to 12 months through long term stability study.
Trade Name
SP1-EGF
(INCI name : processing)
Source
E. coli
Purity
>95% (HPLC)
Amino Acid 99
Molecular Weight
11.9 kDa
pH value
8.0
Activity unit
>5 X 105 IU/mg
Formulation
Lyophilized or Solution stock
Concentration
10, 50, 100 ppm
Certificate of Analysis

12. (INCI name : processing) Lyophilized or Solution stock
SP1-FGF1
(Skin Permeable – Acidic Fibroblast Growth Factor)
Characteristics of SP1-FGF1
PTD & FGF1 Fusion Protein
The PTD fusion protein with FGF1 makes skin permeability better than with protein itself so maximize its effects by deep penetration into the cells.
High Purity and High Activity
With purity more than 95% , suitable as a cosmetic
Ingredient and more active than normal FGF1.
Safety and Stability
Free from various microbial contamination and allergen. Suitable for the specification. Through the toxicity test, safety is assured. It is proved to be stable up to 12 months through long term stability study.
Trade Name
SP1-FGF1
(INCI name : processing)
Source
E. coli
Purity
>95% (HPLC)
Amino Acid
171
Molecular Weight
19.9 kDa
pH value
8.0
Activity unit
>1 X 106 IU/mg
Formulation
Lyophilized or Solution stock
Concentration
10, 50, 100 ppm
Certificate of Analysis

13. HPLC Analysis of SP1-EGF HPLC Analysis of SP1-FGF1
Purity of SP1-EGF
SDS-PAGE
Western blot
SP1-EGF
Marker
HPLC Analysis of SP1-EGF
SP1-EGF
98.2%
Purity of SP1-FGF1
SDS-PAGE
Western blot
Marker
SP1-FGF1
HPLC Analysis of SP1-FGF1
SP1-FGF1
96.4%

14. Stability of SP1-EGF Stability of SP1-FGF12 4 3
1. No treatmetnt
2. Normal EGF
3. Store at -60℃
4. Store at 4℃
# Treat Conc. : 100ng/ml
Phospho-EGF receptor (Tyr1068)
SP1-EGF long term storage stability
(12 months)
SP1-FGF1 Lyophilized Product
SP1-FGF1 Solution Product
All stable up to 12 months
Continuous up date
Stability of SP1-FGF1
All stable up to 12 months
Continuous up data
SP1-FGF1 Lyophilized Product
SP1-FGF1 Solution Product
1. No treatment
2. Normal FGF1
3. Store at -60℃
4. Store at 4℃
# Treat Conc. : 100ng/ml
SP1-FGF1 long term storage stability
(12 months) 1 2 4 3
Phospho
FRS2-alpha
(Tyr196)

15. Prototype of SP1-EGF containing cosmetics
Used Cell Line : A431
M : Size ladder
A : Growth Factor Non Treated Group
B : 100ng/ml of Normal EGF Treated Group
0.1 ~ 100ng/ml : Diluted SP1-EGF Containing Cosmetics
Phospho-EGF
receptor
This proves that SP1- EGF contains proper function which can be used as a cosmetics.
Prototype of SP1-FGF1 containing cosmetics
Phospho-FGF1
receptor
Used Cell Line : NIH3T3
M : Size ladder
A : Growth Factor Non Treated Group
B : 100ng/ml of Normal FGF1 Treated Group
0.1 ~ 100ng/ml : Diluted SP1-FGF1 Containing Cosmetics
This proves that SP1- FGF1 contains proper function which can be used as a cosmetics.

16. Clinical evidence of effect for skin renewal
Day 0
Day 28
Dermal Density TEST
Test Sample
Cosmetics containing SP1-EGF and SP1-FGF1
Contents
SP1-EGF : 10ppm / SP1-FGF1 : 10ppm
Subjects Information
Wrinkle degree who SOP standards than Class 4,
36 to 48-year-old(average 43.3±3.1) female subjects 21
Test Period
8 weeks
Test Method
Crow’s feet using a three-dimensional image analysis, dermis density, moisture content measurements facial sag, viscoelastic, and evaluate the effectiveness by improving the product through survey evaluation by subjects

17. Bioceltran Real Effective Cosmeceuticals begin at
MANUFACTURER / SUPPLIER
BIOCELTRAN. INC.
RM# 305, BIO 2BD, Hupyong-Dong, Chuncheon, Gangwon-Do, South Korea
Tel : Fax :
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