Journal of Bacteriology and Virology 2009. Vol. 39, No. 1 p.11 19 Isolation and Identification of Lactic Acid Bacteria Inhibiting the Proliferation of Propionibacterium acnes and Staphylococcus epidermidis Mi-Sun Kang, Hyun-Ju Oh, Hyun-Chul Lee and Jong-Suk Oh * Department of Microbiology, School of Medicine, Chonnam National University, Gwangju, Korea Propionibacterium acnes is the most common causative agent of acne. Staphylococcus epidermidis is another major bacterial strain to be found in acne lesions. Two strains of lactic acid bacteria (LAB) were isolated from normal inhabitants of humans, which inhibited the proliferation of P. acnes and S. epidermidis. The growth of P. acnes and S. epidermidis was decreased by 4-log scales after incubation for 24 h with LAB isolates, whereas the growth rate of selected LAB isolates were not affected by these pathogenic bacteria. This antibacterial activity of LAB isolates was related to lactic acids, hydrogen peroxide and bacteriocin-like compound production. Two LAB isolates efficiently adhered to human keratinocytes HaCaT and were identified by API 50 CHL medium kit and 16S rdna partial sequencing analysis. The similarity of 16S rdna sequences between one isolate and Lactobacillus salivarius subsp. salicinius was 100%, which suggests that they were L. salivarius subsp. salicinius. On the other hand, 16S rdna sequence similarity between the other isolate and Lactobacillus fermentum was 99.04%, which indicates that it was L. fermentum. In conclusion, these results demonstrate that the two LAB strains isolated from human body were identified as L. salivarius subsp. salicinius and L. fermentum, which inhibit the proliferation of P. acnes and S. epidermidis. Key Words: Isolation, Identification, Lactobacillus, Propionibacterium acnes, Staphylococcus epidermidis 서론 여드름은모낭-피지선에서발생하는피부질환이다. 여드름은피지선에서피지분비가증가하거나피지선의모공이좁아지든지막혀서피지가배출되지못함에따라세균이증식하여염증이생기는것이다. 여드름은주로사춘기나이의사람에게서많이발생하는데이는사춘기나이에분비되기시작하는안드로젠이라는남성호르몬때문이다. 안드로젠은피지분비를촉진시키고, 표피의과각화를일으킨다. 피지의분비증가와표피과각화로모낭-피지선에서피지가정체되어모낭이막힘에따라모낭내부가 Propionibacterium acnes를비롯한혐 Received: February 13, 2009/ Revised: March 6, 2009 Accepted: March 11, 2009 * Corresponding author: Jong-Suk Oh, M.D., Ph.D. Department of Microbiology, School of Medicine, Chonnam National University, 5 Hak-Dong, Dong-Gu, Gwangju 501-746, Republic of Korea. Phone: +82-62-220-4134, Fax: +82-62-228-7294, e-mail: joh@chonnam.ac.kr 기성세균이잘자랄수있는환경이된다 (1, 2). 동시에 Staphylococcus epidermidis와같은다른세균들이모낭주위에서여드름과여드름합병증을일으키는데역할을한다 (3). 여드름발생의병리조직학적인기전은 P. acnes 의효소, 사이토카인및보체와중성구, 손상된각질형성세포에서분비된 cytokine 등이염증을일으키는데관여하는것으로알려져있으나 (4), 아직정확한기전에대해서는알려진바없다. P. acnes와 S. epidermidis 등의균들이염증반응을유발하는데주된역할을하게되므로염증성여드름의치료에항생제가사용되고있다 (5). Triclosan, benzoyl peroxide, azelaic acid (6), retinoid, tetracycline, erythromycin, macrolide, clindamycin 등의항생제가사용되고있으나, 부작용이알려져있다. Benzoyl peroxide와 retinoid는피부건조증이나과민증을유발하고 (7), tetracycline, erythromycin, macrolide, clindamycin은항생제에대한내성발생으로인하여지속적인사용이어렵고간독성이심하며, 칸디다증과같은기회감염증이나타날수있다 (8, 9). 또한, triclosan의경 11
12 M-S Kang, et al. 우빛에노출되었을때환경호르몬으로바뀌어심각한환경오염을일으킬수있다. 따라서, 많은연구자들이항균효과가있으면서부작용이없는여드름치료제를개발하려고노력중이다 (10, 11). 유산균은탄수화물을발효하여최종대사산물로유산을생산하는세균을말한다. 유산균은인간과동물의장및질에존재하며 (12, 13), 통상적으로김치또는요구르트등발효식품의제조과정에활용되고있다. 식품에활용되는유산균으로는 Carnobacterium, Enterococcus, Lactobacillus, Lactococcus, Leuconostoc, Oenococcus, Pediococcus, Streptococcus, Tetragenococcus, Vagococcus, Weissella 등이있다 (14, 15). 이와같이이로운방향으로작용하는정상세균중유산막대균인 Lactobacillus는대표적인유산균으로유해균발육을억제하는효과가강하여발효식품과유산균제재, 의약품등으로사용되고있으며, 병원성세균을억제하는 Lactobacillus의분리및동정이연구되고있다 (16~19). 본연구의목적은세균을이용한여드름치료제를연구하기위한첫과정으로, P. acnes와 S. epidermidis의증식을억제하며각질세포에대한부착능이있는유산균을선발하여생화학적성질및 16S rdna partial sequencing analysis에의한동정을실시하고자하였다. 재료및방법공시세균및배양공시세균으로는 P. acnes ATCC11828와 S. epidermidis ATCC 12228 (Rockville, MD, USA) 를이용하였다. P. acnes 는 Actinomyces broth (BBL, Sparks, MD, USA) 에접종하여 37 혐기조건 (85% N 2, 10% H 2, 5% CO 2 ) 에서 1일간배양하였으며, S. epidermidis는 Brain Heart Infusion broth (BHI, Difco, Detroit, MI, USA) 에접종하여 37 호기조건에서 16시간배양하였다. 각각의균은실험에이용하기전에본배지에서 2회계대배양한후실험에이용하였다. 시료채취및유산균의분리인체구강, 장, 질로부터시료를채취하여 0.9% NaCl로계단희석하여유산막대균분리용배지인 Rogosa 우무배지 (Difco) 에접종하고 37 에서 48시간배양한후집락을취하였다. 이것을다시 De Man, Rogosa, Sharpe (MRS) 우무배지 (Difco) 에접종, 배양하여약 3000주의유산균 을분리하였다. Actinomyces 우무배지와 MRS 우무배지가동량섞어진배지표면전체에 P. acnes를도말접종한다음, 분리균주를점점이떨어뜨려 37 배양기에서 24시간배양하여투명환 (clear zone) 을나타내는균주를 1차선발하였다. 선발된균주들은다시 P. acnes와 S. epidermidis와의상호작용을통하여최종적으로유산막대균 2주를선발하여 lactic acid bacteria (LAB) 1, LAB 2로명명하였다. 분리균주 2주는 MRS 액체배지에배양한후글리세롤의최종농도가 20% (w/v) 되도록첨가하여 -80 에냉동보관하면서필요에따라접종, 배양하여실험에사용하였다. 분리유산균의과산화수소생성능측정분리균주의과산화수소의생성능력을보기위하여 0.25 mg/ml TMB (3,3',5,5'-tetramethyl benzidine, Sigma, St. Louis, MO, USA) 0.01 mg/ml peroxidase (Sigma) 가첨가된 MRS 우무배지에각분리균주를 3 μl씩접종하여 37 에서혐기배양한후꺼내어호기상태에서집락의색깔을관찰하여집락의색깔이청색을띠면과산화수소생성양성 (positive) 으로판단하였고색깔에따라감청색은 strongly positive, 청색은 positive, 약청색은 weakly positive, 색깔의변화가없으면음성 (negative) 으로정하였다 (20). 분리유산균의 P. acnes와 S. epidermdis에대한항균작용분리유산균의 P. acnes에대한항균력을알아보기위하여 MRS broth와 Actinomyces broth가동량섞어진배지에접종량이각각 1.0 10 6 CFU가되도록 P. acnes와분리유산균을단독또는병합으로접종하였다. 37 에서 8시간및 24시간혐기배양후배양액을희석하여 MRS 우무배지와 Actinomyces 우무배지상에접종하여 48시간배양한다음, 분리유산균과 P. acnes의생균수를산정하였다. 또한, S. epidermidiis에대한항균력을알아보기위하여 MRS broth와 BHI broth가동량섞어진배지에접종량이각각 1.0 10 6 CFU가되도록 S. epidermidis와분리유산균을단독또는혼합으로접종하였다. 37 에서 8시간및 24시간호기배양후배양액을희석하여 MRS 우무배지와 BHI 우무배지상에접종하여 48시간배양한다음, 분리유산균과 S. epidermidis의생균수를산정하였다.
Lactic Acid Bacteria Inhibiting Propionibacterium acnes and Staphylococcus epidermidis 13 분리유산균배양상청액의 P. acnes와 S. epidermdis 에대한항균작용분리유산균배양상청액의분비물질중에서어떤성분에의한항균효과인지를알아보기위하여분리유산균을 MRS broth에서 24시간배양한다음원심분리 (4,000 rpm, 20 min, 4 ) 한것을여과하여균을완전히제거하였다. 유산에의한항균력인지알아보기위하여상청액에 proteinase K (0.1 mg/ml) 와 catalase (0.5 mg/ml) 를처리하였으며, 과산화수소에의한항균력인지알아보기위하여상청액을 ph 6.3으로중화시키고 proteinase K를처리하였다. 또한, 상청액을 ph 6.3으로중화시키고 catalase를처리하여박테리오신유사물질 (bacteriocin-like compound) 에의한항균력인지알아보고자하였다. P. acnes와 S. epidermidis 배양액을각각흡광도 600 nm에서 0.05가되도록희석하여 96 well plate에 100 μl 접종하고배양상청액을 100 μl 첨가하였으며, 대조군으로는 MRS broth를 100 μl 첨가하였다. 37 에서 24시간혐기및호기배양후 microplate reader 를이용하여 600 nm에서흡광도를측정하였다. 분리유산균의 human keratinocytes HaCaT에대한부착능분리유산균의 human keratinocytes에대한부착능을알아보기위해실험에이용한세포주는각질형성세포주인 HaCaT (epithelial cell line from adult human skin) 으로서, 10% fetal bovine serum (FBS) 과 penicillin (10 U/ml)/streptomycin (10 μg/ml) 을첨가한 Dulbecco's modified Eagle's medium (DMEM, GibcoBRL, Braunschweig, Germany) 배지를사용하여 37, 5% CO 2 배양기에서배양하였다. 분리유산균의부착실험은 Scaletsky 등 (21) 의방법에따라수행하였다. 4-well Lab-Tek II chamber slide system (Nalge Nunc International, Naperville, IL, USA) 에 well당 10 5 세포를 18시간배양한후, PBS로세포를 2회세척하고 multiplicity of infection (MOI) 이 250이되도록분리유산균 (10 9 bacteria/ ml) 을 25 μl 가하여 37, 5% CO 2 배양기에서 30분간배양하였다. PBS로세포를 3회세척한후 methanol로고정하고 30분간 Giemsa 염색한후, PBS로다시세척하여공기중에건조시켜서각각의세포에대한부착력을광학현미경 (BX51, Olympus, Tokyo, Japan) 을이용하여관찰하였으며, 100개의세포에부착한세균의평균개수를 Table 1. Hydrogen peroxide-producing LAB isolates from normal inhabitants of humans Isolates Source H 2 O 2 -productivity LAB 1 Saliva Positive LAB 2 Saliva Strongly positive 측정하였다. 분리유산균의동정 분리유산균의 1차동정법으로는 API 50 CHL kit (BioMérieux, Marcy, l'etoile, France) 를사용하여동정프로그램인 API LAB plus로검사결과를분석하여생리적특성을검토하였으며, 2차동정법으로는 16S rdna 염기서열을분석하여결정하였다. 16S rdna 염기서열분석을하기위하여 Rochelle 등 (22) 의방법으로단일콜로니에서 DNA를분리하였으며, 27F (5'-AGAGTTTGATCMTG- GCTCAG-3') 와 1522R (5'-AAGGAGGTGWTCCARCC-3') primer를사용하여 16S rdna를 PCR 증폭하였다 (23). PCR 산물은 Wizard PCR Preps DNA purification system (Promega, Madison, WI, USA) 을이용하여정제하였으며, 염기서열분석은 BigDye TM Terminator Cycle Sequencing Ready Reaction Kit (Applied Biosystems, Foster City, CA, USA) 와 ABI PRISM TM 310 Genetic Analyzer (Applied Biosystems) 을이용하여염기서열을결정하였다. 얻어진염기서열결과는 NCBI의 GenBank 프로그램을사용하여상동성을조사하였다. 통계처리 통계적처리는 SPSS 통계분석프로그램 (SPSS version 12.0) 을사용하였으며, Mann-Whitney test로분석하였다. 결과 분리유산균의과산화수소생성능력 혐기성세균은과산화수소에예민하므로분리유산균의과산화수소생성능력을측정한결과, LAB 1과 LAB 2 는과산화수소를생성하여감청색의집락을형성하였다 (Table 1). P. acnes 에대한분리유산균의항균력 P. acnes 에대한분리유산균의항균력을보기위하여
14 M-S Kang, et al. P. acnes와분리유산균의혼합배양후생균수검사결과, 24시간배양후두분리유산균모두 P. acnes 생육을유의하게감소시켰다 (p < 0.05). 8시간혼합배양후 P. acnes 에대한분리유산균의항균력은거의없었으나, 24시간배양후단독배양한 P. acnes 생균수는 3.6 10 9 ± 8.0 10 8 CFU/ml이었으며, LAB 1과병합으로배양시에는 P. acnes 생균수는 4.0 10 6 ± 7.0 10 4 CFU/ml으로감소하였다. 또한, LAB 2와병합시 2.0 10 4 ± 3.0 10 3 CFU/ml 으로크게감소하였다. 따라서, 두분리유산균중 P. acnes에대한항균력은 LAB 2가더좋았다. 반면, 두분 리유산균을혼합배양하였을때생균수에대한변화가거의없었다 (Fig. 1). S. epidermidis에대한분리유산균의항균력 S. epidermidis와분리유산균의혼합배양후생균수검사결과, 두분리유산균모두 8시간부터 S. pidermidis 생육을유의하게감소시키기시작하였으며, 24시간후에는 S. pidermidis 생육을 10 4 CFU/ml까지감소시켰다 (p < 0.05). 24시간배양후 S. epidermidis 생균수는 1.7 10 8 Figure 1. Viable cells of P. acnes and the LAB isolates in the mixed cultures over time. Pa, P. acnes. *p < 0.05 for coculture versus monoculture. Values are means ± standard deviations of three independent experiments. Figure 2. Viable cells of S. epidermidis and the LAB isolates in the mixed cultures over time. Se, S. epidermidis. *p < 0.05 for coculture versus monoculture. Values are means ± standard deviations of three independent experiments.
Lactic Acid Bacteria Inhibiting Propionibacterium acnes and Staphylococcus epidermidis 15 Table 2. Antibacterial activity of lactic acid, hydrogen peroxide (H 2 O 2 ) and bacteriocin-like compound (BLC) in cultured supernatants of two LAB isolates against P. acnes and S. epidermidis Isolates Lactic acid a Inhibition (%) * by H 2 O 2 b P. acnes S. epidermidis P. acnes S. epidermidis P. acnes S. epidermidis LAB 1 99.42±0.18 99.13±0.02 19.75±0.23 5.638±3.06 17.75±3.18 10.42±2.08 LAB 2 94.74±5.61 99.01±0.54 17.63±0.47 12.82±8.24 32.15±3.57 15.80±5.56 * Values are means ± standard deviations of three independent experiments. a Inhibition of growth by lactic acid was measured following treatment of the sterilized supernatants with proteinase K (0.1 mg/ml) and catalase (0.5 mg/ml). b H 2 O 2 -dependent activity was evaluated using the neutralized and proteinase K-treated supernatants. c Inhibition of growth by BLC-dependent activity was evaluated using catalase-treated and neutralized supernatants BLC c ± 3.0 10 7 CFU/ml이었으며, LAB 1과병합으로배양시에는 S. epidermidis 생균수는 2.0 10 4 ± 7.0 10 3 CFU/ ml으로감소하였고, LAB 2와의병합시 1.0 10 4 ± 2.0 10 3 CFU/ml으로감소되었다. 따라서, 두분리유산균중 S. epidermidis에대한항균력도 LAB 2가더좋았다. 반면, 두분리유산균모두혼합배양하였을때생균수에대한변화가거의없었다 (Fig. 2). P. acnes 및 S. epidermidis에대한분리유산균배양상청액의항균력 A B P. acnes 및 S. epidermidis에대한분리유산균배양상청액의항균력을조사한결과, 분리유산균 2주모두유산에의하여약 95% 이상항균효과를나타내었으며, 과산화수소및박테리오신유사물질에의해서도 P. acnes의생육을각각 18~19% 와 18~32% 억제하였다 (Table 2). Human keratinocytes HaCaT 에대한유산균의부착력각질형성세포에대한분리유산균의부착능을알아본결과, 분리유산균모두각질형성세포주인 HaCaT 세포에부착이잘되었으며, 1개의 HaCaT 세포당부착된 LAB 1과 LAB 2 평균개수는각각 91.0 ± 9.6.5, 40.0 ± 2.0 개로서, LAB 1이부착력이더우수하였다 (Fig. 3). 유산균의동정탄소원의이용성차이로동정하는생화학적방법인 API 50 CHL kit로 1차동정을한결과, LAB 1은 L. salivarius ( 가능성 99.9%), LAB 2는 Lactobacillus cellobiosus ( 가능성 92.9%), Lactobacillus delbrueckii subsp. delbrueckii ( 가능성 3.1%) 로동정되었다. Figure 3. Microscopic observations and numbers of adhered of LAB isolates to HaCaT cells. A, LAB 1 vs. HaCaT cells; B, LAB 2 vs. HaCaT cells. Magnification, 1000. Values are means ± standard deviations of three independent experiments. 한편 LAB 1는 16S rdna 991 bp를직접염기서열분석하여 (Fig. 4), NCBI의 GenBank 프로그램을사용하여상동성을조사하여본결과, 비교한 991개의염기중 L. salivarius subsp. salicinius JCM 1230 (accession no. AB289295) 과 100% 상동성을보여 L. salivarius subsp. salicinius로동정되었다 (Table 3). LAB 2는 Lactobacillus fermentum ATCC 14931 (accession no. AB017345) 과유사치가 99.04% 로가장높아 L. fermentum으로동정되었다 (Table 4).
16 M-S Kang, et al. Figure 4. Nucleotide sequence of the partially amplified 16S rdna gene from LAB 1 by PCR. Table 3. 16S rdna similarity of LAB 1 to other bacteria Strains L. salivarius subsp. Salicinius JCM 1230 고찰 Similarity (%) Nucleotide differences 100.00 0/991 L. salivarius T 99.90 1/985 L. salivarius subsp. Salivarius ATCC 11741 99.80 2/990 L. aviarius T 94.95 49/970 L. agilis T 92.27 75/970 L. acidipiscis FS60-1 91.68 82/985 L. animalis T 91.58 81/962 여드름은가장흔한피부질환중하나로, 모낭-피지선단위의자기국한성만성염증성질환으로면포, 홍반성구진, 농포등을형성하는것을특징으로하며, 드물게결절혹은가성낭종이발생하고활동성병변의후유증으로소와성혹은비후성반흔을남기기도한다. 여드름의임상양상은비염증성인면포와염증성인표재성병변과심재성병변으로나타나는데, 심재성병변인결절은주로남자에서나타나며결절사이에진피를통한루 (sinus) Table 4. 16S rdna similarity of LAB 2 to other bacteria Strains Similarity (%) Nucleotide differences L. fermentum ATCC 14931 99.04 6/622 L. thermotolerans DSM 14792 93.45 41/626 L. ingluviei LMG 20380 93.45 41/626 L. mucosae DSM 13345 88.42 72/622 L. vaginalis NCTC 12197 87.90 68/562 L. plantarum JCM 1149 86.91 80/611 L. panis DSM 6035 86.85 81/616 L. reuteri DSM 20016 86.80 82/621 L. pentosus JCM 1588 86.76 81/612 L. aviarius subsp. aviarius ATCC 43234 86.43 81/597 L. oralis DSM 4864 86.34 84/615 가형성되어농양및낭종으로커지게되고치유된후에는흔히영구적으로위축성혹은켈로이드성반흔을남긴다. 여드름의원인세균중에서가장대표적이라할수있는 P. acnes는통성혐기성그람양성간균으로호지성인특성이있어서피지분비가많은부위인얼굴, 목, 등, 가슴에호발한다. P. acnes는여드름환자의혈액단핵구로부터 IL-8의분비를유도하거나균으로부터 lipase가
Lactic Acid Bacteria Inhibiting Propionibacterium acnes and Staphylococcus epidermidis 17 분비되어호중구가피부병변에유입된다 (24, 25). 그리고 P. acnes와분비되는 coproporphyrin III에의해각질형성세포가자극되어 IL-1, TNF-α와 GM-CSF와같은 cytokine이분비되어여드름의염증에관여하는것으로알려져있다 (26, 27). 여드름치료에서항생제는효과적이지만, 항생제의장기간사용으로여드름발생에관여하는균주나피부상재균의항생제내성이보고되고있다 (8). 이런항생제내성문제를극복하기위하여대체치료법으로약용식물들이광범위하게연구되어왔다. 최근에는전통약초들을이용한스킨케어화장품을만들어여드름치료를위한새로운기능성분을찾기위한노력이계속되고있다 (10, 28). 또한, 목련줄기껍질에서분리된마그놀롤 (magnolol) 과호노키올 (honokiol) 은피부질환치료제로서이용되어오고있는데. 항염증및항균효과가있다고보고되었다 (29, 30). 병원성세균을억제하는유산균에대한연구는많이되어왔으나, P. acnes에대한항균효과를가진유산균에대한연구는미비한편이다 (31). Smith 등 (32) 은거의모든유산막대균이박테리오신을생성하고, 여성의질내에상재하는유산막대균은과산화수소를분비하여항균력을가지고있으며, 병원체와결합력이높은것으로보고하였다 (33). 혐기성세균은과산화수소에매우예민하므로과산화수소에의하여억제된다 (34, 35). 이러한항균물질을생성하는세균의작용은항생제의부작용과는달리피부환경의균형을파괴시키지않으면서효과를지속적으로발휘할수있어피부건강의유지에중요한역할을할것으로사료된다. 본연구에서분리한유산막대균중에서 L. salivarius subsp. salicinius로동정된 LAB 1 과 L. fermentum으로동정된 LAB 2가과산화수소를생성하는능력이있었으며, LAB 2가과산화수소를더많이생성하였다. P. acnes를단독으로 24시간배양했을때에비해분리유산균과병합시생균수가 10 3 ~10 5 CFU/ml의감소를보였으며, LAB 2가항균력이더우수하였다. 또한, S. epidermidis에대해서도분리유산균과병합시약 10 4 CFU/ml의감소를보였으며, P. acnes에대해서와마찬가지로 LAB 2가항균력이더좋았다. 그리고, 분리유산균의배양상청액또한 P. acnes 와 S. epidermidis의생육을거의억제하였으며, 대부분이유산에의해항균작용을보였으며, 과산화수소및박테리오신유사물질에의해서도어느정도항균력을나타내었다. 각질형성세포에부착이잘일어나는세균은피부의여드름병변에접종하였을때, 피부에서군락화가되기쉬워세포에부착이잘안되는세균에비하여치료효과가더좋을것으로사료된다. 따라서, 본연구에서각질형성세포주인 HaCaT 세포에대한분리유산균의부착정도를본결과, 분리유산균모두부착이잘일어났다. 종래미생물을동정하기위하여이용하던배양이나혈청학적방법에비교하여미생물에존재하는고유한유전자를이용하여동정하는방법은민감도와정확성에있어서월등하기때문에최근에세균으로부터추출한 DNA 에 rrna 유전자 probe를 hybridization시킨후, 제한효소로처리한결과로세균을분류하기시작하였다 (36). 분리균주의 16S rdna 염기서열을비교하면세균이진화과정상가까울수록염기서열은비슷하고진화과정상멀수록염기서열의유사성은감소하게된다. 본연구에서도분리유산균 2주를탄수화물발효로검사하는 API 50 CHL medium kit로동정시험한결과와 16S rdna 유전자와비교한결과가약간의차이가있었는데, 분리유산균 1주는 L. salivarius subsp. salicinius의유전자와유사치가 100% 를보여 L. salivarius subsp. salicinius로동정되었으나, 다른한주는 L. fermentum의유전자와유사치가 99.04% 로서, API 50 CHL medium kit를이용한동정결과인 L. cellobiosus (92.9%) 와는상당한차이가있음을알수있었다. 따라서최종적인세균의동정은유전자분석에의해이루어져야될것으로사료된다. 본연구에서는 L. salivarius subsp. salicinius와 L. fermentum으로동정된분리유산균들이 P. acnes와 S. epidermidis를억제하는효과가있었고, 각질형성세포인 HaCaT 세포에대한부착력이좋았으며, 과산화수소생성능력이우수한 L. fermentum이 P. acnes와 S. epidermdis에대해서도가장억제력이우수하였다. 향후이연구의결과가사람의여드름병변에서도임상적으로검증된다면분리유산균의여드름예방과치료를위한프로바이오틱으로개발하여건강의증진에기여할수있을것으로생각된다. 또한, P. acnes와 S. epidermidis에대한분리유산균의억제기전에대한추가연구가필요할것으로사료된다. 참고문헌 1) Harper JC. An update on the pathogenesis and management of
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