산국 (Chrysanthemum boreale) 휘발성 추출액의 항균 및 항진균활성

차정단,김태영,우원홍,정규용,김강주,길봉섭 원광대학교 생체재료·매식연구소 1999 원광생체재료·매식 Vol.8 No.1

To investigate the antimicrobial and antifungal activity of Chrysanthemum boreale(C. boreale) essential oil, the growth inhibition against Aspergillus nidulans, Fusarium oxysporum, Alternaria mali, Staphylococcus aureus, Streotococcus pyogenes, Streptococcus mutans and Escherichia coli in the presence of the essental oil was monitored by agar dilution method. The antifungal effect of C. boreale on Aspergillus nidulans, Fusarium oxysporum and Alternaria mali as affected at the concentration of 400 ppm. The antibacterial effect of C. boreale on staphylococcus aureus, streptococcus pyogenes and Streptococcus mutans was affected at the concentration of 0.4 μl, while growth of E. coli was not inhibited at the concentration of essential oil. The results of this experiment on microorganisms indicated that naturally occuring chemical substances from C. boreale might have biologically toxic activities, that is, allelopathic potentials to the microorganisms.

산국 (Chrysanthemum boreale) 휘발성 추출액 및 EDTA가 미생물 및 인체 치은 섬유아세포에 미치는 영향

차정단,김태영,우원홍,전병훈,김해경,유용욱,김강주,길봉섭 원광대학교 생체재료·매식연구소 2000 원광생체재료·매식 Vol.9 No.1

Chrysanthemum boreale (C. boreale) has been used for the treatment of furuncles, carbuncles, sores, conjunctivitis, headache, vertigo and swollen throats as a folk medicine in Korea, but there is little report about experiment evidence of antimicrobial activity. To investigate the antifungal and antibacterial activity of Chrysanthemum boreale(C. boreale) volatile substance and EDTA, the growth inhibition against Aspergillus nidulans (A. nidulans), Fusarium oxysporum (F. oxysporum), Alternaria mali(Al. mali), Candida albicans (C. albicans), staphylococcus aureus (S. aureus), Streptococcus pyogenes(S. pyogenes), streptococcus mutans (S. mutans), Escherichia coli(E. coli), Helicobacter pylori (H. pylori) and Porphyromonas gingivalis (P. gingivalis) was monitored by agar dilution and broth microdilution method. The antifungal effect of C. boreale on A. nidulans, F. oxysporum, Al. mali and C. albicans was shown at the concentration of 0.04%. The antibaterial effect of C. boreale on S. aureus, S. pyogenes, S. mutans and H. pylori was shown at the concentration of 0.04%. However, the growth was inhibited at the concentration of 0.04% irrespective of the presence of EDTA 0.125% was much higher than higher than that of several concentration of volatile substance. The minimum inhibitory concentrations of volatile substance were 0.16% against S. aureus, 0.04% against S. pyogenes, 0.02% against S. mutans, 0.40% against E. coli, and 0.18% against C. albicans. The growth of human gingival fibroblasts was not affected by C. boreale volatile substances at the concentration of 0.04%. These results indicate that volatile substance from C. boreale may have biologically toxic activities to the microorganisms.

Berberine의 항생제 내성 확색포도구균에 대한 항미생물 효과

김강주,차정단,김해경,유현희,유용욱,이동근 대한악안면성형재건외과학회 2002 Maxillofacial Plastic Reconstructive Surgery Vol.24 No.2

Antibiotic-resistant Staphylococcus aureus (S. aureus) emerged in many parts of the world and increased complex clinical problems. Therefore, new agents are needed to treat the antibiotic-resistant S. aureus. Some natural products are candidates of new antibiotic substances. Previous reports showed that the extract of Coptidis Rhizoma and Phellodendri Cortex have antimicrobial effects. The main antibacterial substance of Coptidis Rhizoma and Phellodendri Cortex is berberine. In this study, we examined the antimicrobial activity of berberine to antibiotic-resistant S. aureus. We isolated the 13 strains of antibiotic-resistant S. aureus from the oral cavity of healthy children in a preschool, and investigated the antimicrobial activity of berberine. We examined the effects of berberine on the level of resistance to antibiotic-resistant S. aureus. We observed that the invasion of S. aureus into human gingival fibroblasts was blocked by berberine. Berberine inhibited the growth of antibiotic-resistant S. aureus as a dosw dependent maner. Minimum inhibitory concentration (MIC) of berberine against antibiotic-resistant S. aureus was 50∼100㎍/㎖. The inhibition of S. aureus invasion was depended on the concentration of berberine in human gingival fibroblasts. The MIC of ampicillin against antibiotic-resistant S. aureus was 32㎍/㎖. However, the MIC of ampicillin against antibiotic-resistant S. aureus in the presence of 50㎍/㎖ berberine was 1㎍/㎖. These data indicate that the combination with berberine and β-lactam antibiotics resulted in the synergistic reaction. These results suggest that the berberine may have the antimicrobial activities to antibiotic-resistnat S. aureus, and the mechanism may be the inhibition of growth and intracellular invasion, and alteration of antibiotic-resistance level of antibiotic-resistant S. aureus.

Berberine의 항생제 내성 황색포도구균에 대한 항미생물 효과

김해경,차정단,유용욱,김강주,이동근 원광대학교 생체재료·매식연구소 2000 원광생체재료·매식 Vol.9 No.1

Antibiotic-resistant Staphylococcus aureus (S. aureus) emerged in many parts of the world and increased complex clinical problems. Therefore, new agents are needed to treat to antibiotic-resistant S. aureus. Some natural products are candidates of new antibotic substances. Previous reports showed that the extract of Coptidis Rhizoma and Phellodendri Cortex have antimicrobial effects. The main antibacterial substance of Coptidis Rhizoma and Phellodendri Cortex is berberine. In this study, we examined the antimicrobial activity of berberine to antibiotic-resistant S. aureus. We isolated the 13 strains of antibiotic-resistant S. aureus from the oral cavity of healthy children in a preschool, and investigated the antimicrobial activity of berberine. We examined the effects of berberine on the level of resistance to antibiotic-resistant S. aureus. We observed that the invasion of S. aureus into human gingival fibroblasts was blocked by berberine. Berberine inhibited the growth of antibiotic-resistant. S. aureus as a dose dependent maner. Minimum inhibition concentration (MIC) of berberine against antibiotic-resistant S. aureus was 50-100 ㎍/ml. The inhibition of S. aureus invasion was depended on the concentration of berberine in human gingival fibroblasts. The MIC of ampicillin against antibiotic-resistant S. aureus was 32 ㎍/ml. However, the MIC of ampicillin against antibiotic-resitant S. aureus in the presence of 50 ㎍/ml berberine was 1 ㎍/ml. These data indicate that the combination with berberine and β-lactam antibiotics resulted in the synergistic reaction. These results suggest that the berberine may have the antimicrobial activities to antibiotic-resistant. S. aureus, and the mechanism may be the inhibition of growth and intracellular invasion. and alteration of antibiotic-resistance level of antibiotic-resistant S. aureus.

급성 구강감염부위에서 Staphylococcus lugunensis 의 특성

유용욱,이미성,차정단,김기경,신상희,문상은,김강주 원광대학교 생명공학연구소 1999 생명공학연구소보 Vol.6 No.1

Staphylococcus lugdunensis (S. lugdunensis) is a newly identified pathogenic species of coagulase negative staphylococci and an occasional but not rare cause of severe infections, such as infective endocarditis after dental extraction, bacteremia, osteomyelitis, peritonitis, and soft tissue infections. As antibiotic use increased, resistance rapidly developed. Some strains have plasmids related to antibiotic resistance. To characterize S. lugdunensis in acute oral infection, S. lugdunensis was isolated from the patients with acute oral abscess, osteomyelitis, and normal persons. Antibiotic resistance, in vitro cellular toxicity, in vivo virulence, δ-like hemolysin activity, and synergistic hemolysis on sheep blood agar plates were investigated. The dot blot analysis and Southern blot analysis of staphylococcal DNA was performed with δ-hemolysin gene probe of Staphylococcus aureus. (S. aureus) Staphylococcal DNA was cloned, nucleotide sequence was analysed, and homology was compared with other sequence in Gene Bank. S. aureus, S. lugdunensis, Staphylococcus cohnii (S. cohnii), and other coagulase negative staphylococci (CNS) were isolated from the patients with acute oral infection. The isolation ratio of S. lugdunensis in the patients with infection was higher than that of healthy persons, but the isolation ratio of S. aureus in the patients with infection was similar to that of healthy persons. S. lugdunensis from the patients with acute oral infection showed the resistance to penicillin, methicillin, cephalothin, and clindamycin. S. lugdunensis in the patients had cellular toxicity in vitro and virulence in vivo. All strains of S. lugdunensis had δ-like hemolysin activity against rabbit erythrocytes. Four of the six strains of S. lugdunensis gave synergistic hemolysis with S. aureus on sheep blood agar plates. In dot blot analysis, all strains of S. lugdunensis showed the positive reaction with the probe of δ-hemolysin gene in S. aureus, but a 7.3 kb HindⅢ fragment was observed in the DNA of S. lugdunensis that gave synergistic hemolysis in a Southern blot analysis. The molecular size of partially purified δ-hemolysin was about 50 kd. The cloned fragments from the chromosomal DNA of S. lugdunensis showed the partial homology with the insulin receptor-related and dopamine receptor of humans. These results suggest that S. lugdunensis might be an important pathogen in acute oral infection and show some homology with eukaryotes.

Intracellular Invasion of Staphylococcus aureus against Human Gingival Fibroblasts : A Potential Pathogenic Mechanism in Oral Infection

Cha, Joung-Dan,Jung, Kyu-Yong,Kim, Hae-Kyoung,You, Yong-Ouk,Kim, Kang-Ju 원광대학교 생체재료·매식연구소 2000 원광생체재료·매식 Vol.9 No.1

Staphylococcus aureus(S. aureus) is one of the important pathogens in acute oral infection, but the pathogenic mechanism of S. aureus is not fully understood. In our previous study, interaction of staphylococcal protein A and cytoskeletal actin filaments is involved in the S. aureus invasion of cultured KB cells, an established human epidermoid carcinoma cell line. To investigate the pathogenic mechanism of S. aureus in oral infection, the invasion of S. aureus in human gingival fibroblasts(HGFs) was studied. Invasion of S. aureus ATCC 25923 and OPT 2 was found to be time-dependent (0-120 min). That was increased linearly when an increasing number of bacteria were added to media (10^2 - 10^7 cfu/mL/well). The invasion of wild type stain (OPT 2) isolated from the patient with acute oral and maxillofacial infection was higher than that of standard strain (ATCC 25923). However the invasion of S. aureus Wood 46, which is deficient in protein A, was notably lower than those of the strains that have protein A (ATCC 25923 and OPT 2). Cytochalasin D that inhibits the organization of actin filaments inhibited the invasion of S. aureus (ATCC 25923 and OPT 2) in HGFs. These results suggest that pathogenic mechanism of S. aureus in oral infection may be associated with intracellular invasion, and interaction of staphylococcal protein A and cytoskeltal actin filament may be involved in the invasion to HGFs.

In vitro Virulence of Cultured Supernatant in Porphyromonas gingivalis W50 under Hemin and Menadione Limited Culture Condition

Moon, Sang-Eun,Cha, Joung-Dan,Kim, Ki-Gyung,Lee, Hyun-Ok,Kim, Kang-Ju 원광대학교 생체재료·매식연구소 1997 원광생체재료·매식 Vol.6 No.1

In order to investigate the change of virulence in P. gingivalis W50, in vitro virulence was measured by the change of cell activity in mouse 3T3 cell. The cell activity of cultured supernatant of P. gingivalis W50 in the presence of hemin was lower than that in the obsence of hemin and menadione, but there was no change according to the presence of menadione. These results suggest that the virulence of cultured supernatant in P. gingivalis W50 might be increased by hemin.

Nucleotide sequence of Staphylococcus lugdunensis agr gene cluster, and comparison of its gene products with others

Lee, Mi-Sung,Cha, Joung-Dan,You, Yong-Ouk,Yoon, Ki-Hong,Kim, Kang-Ju 원광대학교 생체재료·매식연구소 2000 원광생체재료·매식 Vol.9 No.1

Staphylococcus lugdunensis, a recently delineated coagulase negative Staphylococci, was known to produce δ-hemolysin. It causes oral infection and severe infection such as acute bacterial endocarditis. Arg gene cluster encoding proteins of a accessory gene regulator was cloned form S. lugdunensis. The agrgene cluster of S. lugdunensis was composed of agrA, agrB, agrC, agrD, and orf5. Nucleotide sequences of agrA, agrB and orf5 was completely determined, indentifying that they encode proteins of 239, 430 and 262 amino acids, respectively. The deduced amino acid sequence of the S. lugdunensis AgrA shows high homology with the S. aureus AgrA (86.4%) and S. epidermidis AgrA (82.6%) S. lugdunensis AgrB is also homologous to the S. aureus AgrB(51.1%), but nor homologous to S. epidermidis AgrB. In addition, the S. lugdunensis ORF5 shows high homology with the S. aureus (60.6%) on the basis of their deduced amino acid sequences. The hld gene coding for δ-hemolysin is located between agrD and orf5 in S. aureus, but hld gene was not found around S. lugdunensis orf5 gene even though s. lugdunensis arg genes and their products are similar to those of S. aureus. These results suggest that the nucletide sequence of S. lugdunensis gene is highly homologus with that of S. aureus agr gene.

Detection and Partial Sequence Analysis of Helicobacter pylori in Gingival Crevicular Fluid

Yang, Chun-Ho,Cha, Joung-Dan,Lee, Mi-Sung,You, Yong-Ouk,Yoon, Ki-Hong,Kim, Kang-Ju 원광대학교 생체재료·매식연구소 2000 원광생체재료·매식 Vol.9 No.1

Many researchers hypothesized that Helicobacter pylori can be transmitted via oral cavity, and oral cavity may be a reservoir. To investigate the characteristics of H. pylori in oral cavity, we amplified and sequenced the urease asubunit of H. pylori in gingival crevicular fluid, and compared the genetic and amino acid homology of H. pylori among human isolates. Oral samples were taken from the patients with gastric disease and detection of H. pylori was performed by the polymerase chain reaction (PCR) with the primer of specific urease of H. pylori 26695 that was isolated form stomach. The amplified DNA fragments were sequenced by an automated DNA sequencer. DNA of H. pylori was detected in 2 out of 42 patients(4.8%) by PCR. The sequenced PCR products of H. pylori urease asubunit showed the genetic heterogeneity with that of H. pylori 26695 which was isolated from stomach, and the deduced amino acid sequence of H. pylori urease asubunit isolated form oral cavities was completely consistent with the asubunit of specific urease of H. pylori 26695 and J99, stomach isolates. These results suggest that H. pylori in gingival crevicular fluid showed the genetic hetergeneity and amino acid homology.