2011

NoA (M3_ )
20400130
20600098
20800647


Abstract ............................................................................................................................................................................................ 1
Introduction ................................................................................................................................................................................... 2
Method and Material ................................................................................................................................................................ 3
.......................................................................................................................................................... 3
...................................................................................................................................................................... 3
...................................................................................................................................................... 3
............................................................................................................................................................................... 3
XTT assay .............................................................................................................................................................................. 3
............................................................................................................................................................................... 3
.......................................................................................................................................................................... 4
............................................................................................................................................................................... 4
.................................................................................................................................................................... 4
MIC ............................................................................................................................................................................... 4
.......................................................................................................................................... 5
.................................................................................................................................................................... 5
Result and Discussion ................................................................................................................................................................ 6
CCD-968sk ........................................................................................... 6
................................................................................................................................. 6
............................................................................................................................................... 6
.................................................................................................................................................................... 7
................................................................................................................................................. 8
Conclusion ...................................................................................................................................................................................... 9
Reference ...................................................................................................................................................................................... 10

Abstract

. ,
(Propionibacterium acnes) .
, , P. acnes .
(Korean Mistletoe
Lectin) .
(Mannose Binding Lectin) .
, .

P. acne .

Introduction
.
, , ,
.
. ,
.
, , , .

A , , ,
(benzoyl peroxide)
, ,
A .
(Propionibacterium acnes)
, ,
,
.
(complement)
, .
()
,
.
,
, .
(mistletoe, Viscum album)
, , ,
.
,
,
.
adjuvant . (1)
(mannose binding lectin; MBL) (Lectin
pathway), .
.
(pearl) 20

.
.
2

Method and Material

, .
800~1200
(2).

10 (w/v) PBS (Lonza, Switzerland)
4 16 4, 10,000g 20 .
ammonium sulfate 70%
. 4 16 4, 10,000g 20
. 50ml PBS
2 4, 10,000g 20 .
0.45um membrane filter (Whatman, England) . (3)

anti-treated Sepharose column loading column
5 PBS . Column 0.1M
lactose PBS PBS lactose .
(4)

CCD-986sk (KCLB 21947) 10%
FBS (Fetal Bovine Serum) (Gibco, USA), 1% Penicillin-Streptomycin (Gibco, USA) DMEM
(Dulbecco's Modified Eagle's Medium) (Gibco, USA) . 37
5% .
(II) ( 11-1470000-000863-01) (5).
XTT assay
CCD-986sk 96 well microtitre plate 5103cell/well
. 500ug/ml 15.626ug/ml 2
100ul duplicate . 48 1mg/ml XTT solution (Sigma, USA)
50ul 4 490nm .

(Propionibacterium acnes)
(KCTC 3314) . P. acnes 10ml (Actinomyces
3

broth) (BD, USA) 3 1 . 1 10%

CO2 . 48 0.2%
(formaldehyde) (Sigma, USA) 24 .
4,000rpm 20 PBS 3 .
.
3 -20 .

6 , FCA (Freund's
Complete Adjuvant) (Sigma, USA) / FIA (Freund's Incomplete Adjuvant) (Sigma, USA) 1:1
1 3 (subcutaneous injection) .

1 2000rcf 10
-20

Time-resolved fluorescence Immunoassay (TR-FIA) (titer) .
Kensner (6) . P. acnes ( 10ug/ml) 96well
microtitre plate 0.1ml 37 2 100ul 100ul
(normal IgG ) 37 2 . , 5~6
(europium)
anti-mouse IgG-Eu particle conjugate 1:5,000 37 2
. , 5~6 200ul (enhancement solution)
(Perkin Elmer, USA) 5 DELFIA Reader (Perkin Elmer, USA)
. (7).
MIC
(minimal inhibitory concentration,
MIC) Clinical Laboratory Standards Institute (CLSI) broth dilution procedure
. 1 mg/ml RCM (reinforced
clostridial medium) (BD, USA) 1/2 . P.

acnes 96-well microtitre plate 105CFU/ml 37 3

. 540nm broth
MIC . tetracycline positive control
(8) (9) (10) (11).


P. acnes P. acnes
. 100, 200, 400, 800ug/ml PBS 30ul 6mm
(Whatman, UK) 37 48 . clear zone
.

.
(BD, USA) P. acnes (Brucella Broth) (BD,
USA) 100ml shaking incubator (Vision Scientific, Korea) , spectrophotometer
(Biochrom, UK) 600nm 0.6 .
30 , 10 109
106, 107, 108, 109
.

Result and Discussion

CCD-968sk
CCD-986sk
XTT assay . XTT assay 250ug/ml
. 250ug/ml .
3

OD value (490nm)

2.5
2
1.5
1
0.5
0
0

100

200

300

400

500

600

(ug/mL)
< 1>

tetracycline .
tetracycline 0.077 0.046 0.102 0.039
.
< 1>
MIC (ug/ml) (n=10)

0.102 0.039

Tetracycline

0.077 0.046

6mm disc clear zone .

P. acnes
clear zone . 200ug/ml clear zone
.

< 2> (paper disc = 6 mm)

P. acnes

0 ug/ml

100 ug/ml

200 ug/ml

400 ug/ml

800 ug/ml

6.5

P. acnes .
(Europium, Eu) tracer TR-FIA ( 2). 3
(Korean Mistletoe Lectin, KMLC) adjuvant
. FCA FIA
P. acnes
. P. acnes
.

< 2>
600000

500000
400000
300000
200000
100000
0
Control

P. acnes

KMLC

< 3>

FCA

FIA



. P. acnes
4 , 1:200,000 .
P. acnes 5 .
90
80
(%)

70
60
50
40
30
20
10
0

< 4> P. acnes
100
90
(%)

80
70
60
50
40
30
20
10
0

+KMLC

+MBL

+KMLC

+KMLC

+MBL

+MBL
+

< 5> P. acnes

8

Conclusion

200ug/ml P. acnes
. .
250ug/ml
P. acnes TR-FIA
adjuvant
. P. acnes

P. acnes .
P. acnes .
.
.


.

Reference
1. , . . s.l. : , 2004.
2. , . . 1010635440000 , 09 01, 2011.
3. Isolation and Characterization of Two Korean Mistletoe Lectins. Kang, Tae Bong, et al. 6, 11
2007, Journal of Biochemistry and Molecular Biology, Vol. 40, pp. 959-965.
4. Cellular and humoral adjuvant activity of lectins isolated from Korean mistletoe (Viscum album

colaratum). Yoon, Taek Joon, et al. 2001, International Immunopharmacology, Vol. 1, pp. 881-889.
5. . (II). s.l. : ,
2005. . 11-1470000-000863-01.
6. Validations of time-resolved fluoroimmunoassays for urinary estrone 3-glucuronide and

pregnanediol 3-glucuronide. Kesner, James S., et al. 3, 3 1994, Steroids, Vol. 59, pp. 205-211.
7. HeterologousELISA . Park, Sung Min, et al. 3, 2010,
Reproductive & Developmental Biology, Vol. 34, pp. 161-167.
8. Propionibacterium acnes . Weon, Jin Bae,
Ahn, Juhee and Ma, Choong Je. 1, 2011, , Vol. 42, pp. 98-101.
9. Propionibacterium acnes . ,
, et al. 3, s.l. : , 6 1995, , Vol. 33, pp. 437-444.
10. , . Propionibacterium acne
. . s.l. : , 2 2010.
11. (Dryopteris crassirhizoma Nakai) Propionibacterium acnes .
Kim, Hyun Ju, et al. 3, 9 2006, Journal of the Society of Cosmetic Scientists of Korea, Vol. 32, pp.
201-208.

10

1,500,000

1,279,800

500,000

407,000

201,000

112,200

()

4,000,000

11

2011 (ACE)

12