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圖 

 

 

Coffea arabica and its hydrolysates

PG DC 100 CA CAH1 CAH2 CAH3 CAH4

 

Michelia alba and its hydrolysates

(1000 μg/mL)

 

Morinda citrifolia and its Hydrolysates

***

*** ***

***

 

 

Caffeic acid & Chlorogenic acid

PG

Fig.1 The inhibition of Coffea arabica, Michelia alba and Morinda citrifolia extract and their hydrolysates on collagenase activity.

(a) Coffea arabica and its hydrolysates 1000 μg/mL (b) Coffea arabica 50-1000 μg/mL (c) Michelia alba and its hydrolysates 1000 μg/mL (d) Michelia alba 50-1000 μg/mL (e) Morinda citrifolia and its hydrolystaes 1000 μg/mL (f) Coffea

arabica (1000 μg/mL), chlorogenic acid and caffeic acid (100 μM)

(Hydrolysate 1: hydrolyzed by 1.2 N HCl for 30 min; hydrolysate 2: 1.2 N HCl for

60 min; hydrolysate 3: 2.4 N HCl for 30min; hydrolysate 4: 2.4 N HCl for 60min)

(*, P< 0.05; **, P< 0.01; ***, P< 0.001)

caffea arabica

Fig. 2  The inhibition (%) of Coffea arabica extract and its hydrolysates on the activity of bacterial collagenase.

Michelia alba

Fig. 3 The inhibition (%) of Michelia alba extract and its hydrolysates on the activity of bacterial collagenase.

Michelia alba

Fig. 4 The inhibition (%) of Michelia alba extract and its hydrolysates on the activity of porcine elastase. (**, P< 0.01; ***, P< 0.001)

 

Coffea arabica

(μg/mL)

0 1 10 100 1000

Cell viability (% of control)

0

Fig. 5 Cell viability (%) of Coffea arabica extract and its hydrolysates on human foreskin fibroblast. (*, P< 0.05; **, P< 0.01; ***, P< 0.001)

Michelia alba

(μg/mL)

0 1 10 100 1000

Cell viability (% of control)

0

Fig. 6 Cell viability (%) of Michelia alba extract and its hydrolysates on human foreskin fibroblast. (*, P< 0.05; **, P< 0.01)

Chorogenic acid & caffeic acid

0 5 10 20 40 80 (μM)

Cell viability (% of control)

0

140 Chorogenic acid

Caffeic acid

   

Fig. 7 Cell viability (%) of chlorogenic acid and caffeic acid on human foreskin fibroblast.

UV dose

(mJ/cm2 )

0 20 40 60 80 100

cell viability ( % of control )

0 20 40 60 80 100

***

Fig. 8 Cell viability (%) of various dose UVB irradiation on human foreskin fibroblast. (***, P< 0.001)

         

UV-induced Expression of MMPs and Procollagen

(mJ/cm )

0 20 40 60 80

Relative density ( of control)

0

Type I procollagen

2

   

 

   

UV-induced expression of MAP Kinase

( mJ/cm )

0 20 40 60 80

Relative density ( of control)

0

Fig. 9 UVB-induced expression of MMP-1, 3, 9, type I procollagen and MAP kinase. (**, P< 0.01; ***, P< 0.001)

 

 

 

(b)

 

Relative density ( of control)

0.0

Type I procollagen

** ** **

Fig. 10 Effect of Coffea arabica extract on the UVB-induced expression of MMP-1, 3 and 9 and type I procollagen in human fibroblast.

(*, P< 0.05; **, P< 0.01; ***, P< 0.001)

 

Coffea arabica

(μg/mL)

control UV 5 10 25

Relative density ( of control )

0.0 0.5 1.0 1.5 2.0 2.5

pJNK pERK pp38

* **

***

***

** ***

Fig. 11 Effect of Coffea arabica extract on the UVB-induced expression of MAP kinases in human fibroblast. (*, P< 0.05; **, P< 0.01; ***, P<

0.001).

Caffeic acid and Chlorogenic acid

( μM ) control UV 5 10 50 5 10 50

Relative density (of control)

0 1 2 3 4

MMP-1 MMP-3 MMP-9 Type I procollagen

Caffeic acid Chlorogenic acid

** **

*

*

* *

*

   

Fig. 12 Effect of caffeic acid and chlorogenic acid on the UVB-induced expression of MMP-1, 3, 9 and type I procollagen in human fibroblast. (*, P< 0.05; **, P< 0.01).

( μg/mL )

control UV 5 10 25

Rel a ti ve density ( of control )

0

Type I procollagen

Michelia alba

*

Fig. 13 Effect of Michelia alba extract on the UVB-induced expression of MMP-1, 3, 9 and type I procollagen in human fibroblast. (*, P< 0.05; **, P< 0.01; ***, P< 0.001).

 

 

 

Michelia alba

( μg/mL )

control UV 5 10 25

Relative density ( of control )

0 1 2 3 4

5 pJNK

pERK pp38

* **

**

**

   

Fig. 14 Effect of Michelia albaextract on the UVB-induced expression of MAP kinases in human fibroblast. ( *, P< 0.05; **, P< 0.01).

Fig. 15 The primary skin irritation test of Coffea arabica extract in the rabbit. (a) Before treating with Coffea arabica extract (b) After applying to skin with 0.1 g and 0.5 g of Coffea arabica extract (c) After a 24 hour exposure (d) After a 72 hour exposure.

Fig. 16 The primary skin irritation test of Michelia alba extract in the rabbit. (a) Before treating with Michelia alba (b) After applying to skin with 0.1 g and 0.5 g of Michelia alba extract (c) After a 24 hour exposure (d) After a 72 hour exposure.

Fig. 17 The Draize eye test of Coffea arabica extract in the rabbit.

Fig. 18 The Draize eye test of Michelia alba extract in the rabbit.

Table 1 The inhibition rate of Coffea arabica and Michelia alba extract on bacterial collagenase by gelatin digestion assay

concentration (μg/mL)

Coffea arabica Michelia alba

50 52.7 ± 0.12 45.9 ± 0.52 100 97.7 ± 0.11 40.0 ± 0.54 500 101.7 ± 0.01 101.2 ± 0.03 1000 100.1 ± 0.03 97.6 ± 0.35

Table 2 The inhibition rate of Coffea arabica, Michelia alba and Morinda

citrifolia extract and their hydrolysates on bacterial collagenase by gelatin

digestion assay

Coffea arabica Michelia alba Morinda citrifolia

extract 100.0 ± 0.05 100.1 ± 0.03 74.3 ± 1.04 H1 99.5 ± 0.06 99.3 ± 0.13 -2.7 ± 2.11 H2 96.7 ± 0.08 99.1 ± 0.14 69.9 ± 1.26 H3 97.1 ± 0.07 99.2 ± 0.12 60.5 ± 17.80 H4 95.1 ± 0.23 93.7 ± 0.40 -3.3 ± 0.97 Hydrolysate 1: 1.2 N, 30 min; hydrolysate 2: 1.2 N, 60 min; hydrolysate 3:

2.4 N, 30min; hydrolysate 4: 2.4 N, 60min

Table 3 The inhibition rate of Coffea arabica extract and its hydrolysates on bacterial collagenase using fluorometric assay

concentration Hydrolysate 1: 1.2 N, 30 min; hydrolysate 2: 1.2 N, 60 min; hydrolysate 3:

2.4 N, 30min; hydrolysate 4: 2.4 N, 60min

Plant sample %

Table 4 The inhibition rate of Michelia alba extract on bacterial collagenase using fluorometric assay

concentration (μg/mL)

Michelia alba

extract H1 H2 H3 H4 10 92.7 ± 0.72 33.6 ± 1.46 36.2 ± 16.7 49.1 ± 5.95 47.1 ± 2.00

50 94.8 ± 0.53 55.6 ± 0.79 68.1 ± 2.23 74.3 ± 5.83 68.9 ± 0.81 100 99.9 ± 0.40 82.3 ± 0.23 91.0 ± 0.90 92.9 ± 0.15 83.1 ± 1.86 500 100.1 ± 0.37 96.0 ± 0.08 99.0 ± 0.07 96.6 ± 0.47 98.7 ± 1.14

Table 5 The inhibition rate of Michelia alba extract and its hydrolysates on porcine elastase

concentration (μg/mL)

Michelia alba

extract H1 H2 H3 H4 50

17.2 ± 0.7 15.5 ± 1.4 7.8 ± 6.7 9.9 ± 3.0 6.4 ± 1.5

100

17.4 ± 0.5 16.2 ± 0.0 8.4 ± 2.2 6.4 ± 3.8 7.4 ± 0.8

500

17.0 ± 0.4 15.0 ± 0.2 6.6 ± 0.9 10.5 ± 0.2 11.4 ± 0.2

1000

29.3 ± 0.3 15.0 ± 0.1 24.8 ± 0.2 12.7 ± 0.5 12.4 ± 0.1

Table 6 The scores of primary skin irritation test of Coffea arabica extract at low dose (0.1 g) in the rabbit

Rat 1 2 3 4 Mean± SD

Time ( h) intact abraded intact abraded intact abraded intact abraded intact abraded 24 0 2 0 0 0 3 0 0 0.00 ± 0.00 1.25 ± 1.30 48 0 0 0 0 0 1 0 0 0.00 ± 0.00 0.25 ± 0.43

     

       

Table 7 The scores of primary skin irritation test of Coffea arabica extract at high dose (0.5 g) in the rabbit

Rat 1 2 3 4 Mean± SD

Time ( h) intact abraded intact abraded intact abraded intact abraded intact abraded 24 0 3 0 0 0 5 0 0 0.00 ± 0.00 2.00 ± 2.12 48 0 1 0 0 0 1 0 0 0.00 ± 0.00 0.50 ± 0.50

 

Table 8 The scores of primary skin irritation test of Michelia alba extract at low dose (0.1 g) in the rabbit

Rat 1 2 3 4 Mean± SD

Time ( h) intact abraded intact abraded intact abraded intact abraded intact abraded 24 0 0 0 0 0 0 0 0 0.00 ± 0.00 0.00 ± 0.00 48 0 0 0 0 0 0 0 0 0.00 ± 0.00 0.00 ± 0.00

Table 9 The scores of primary skin irritation test of Michelia alba extract at high dose (0.5 g) in the rabbit

Rat 1 2 3 4 Mean± SD

Time ( h) intact abraded intact abraded intact abraded intact abraded intact abraded 24 0 0 0 1 0 1 0 0 0.00 ± 0.00 0.50 ± 0.50 48 0 0 0 1 0 0 0 1 0.00 ± 0.00 0.50 ± 0.50

 

Table 10 The scores of Draize eye test of Coffea arabica Rat

Time (h) 1 2 3 Mean± SD

1 0 0 0 0.00 ± 0.00 24 0 0 0 0.00 ± 0.00 48 0 0 0 0.00 ± 0.00 72 0 0 0 0.00 ± 0.00

 

Table 11 The scores of Draize eye test of Michelia alba Rat

Time (h) 1 2 3 4 Mean± SD

1 0 0 0 0 0.00 ± 0.00 24 0 0 0 0 0.00 ± 0.00 48 0 0 0 0 0.00 ± 0.00 72 0 0 0 0 0.00 ± 0.00  

 

附錄一 Draize evaluation of dermal reactions

SKIN REACTIONS SCORE

Erythema and Eschar Formation (Most severely affected area graded):

No erythema 0

Very slight erythema (barely perceptible) 1

Well-defined erythema 2

Well-defined erythema 2

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