椰子油对变形链球菌抑菌作用的体外研究

目的研究椰子油对变形链球菌的生长抑制作用,通过观察其对生物膜活性、产酸及粘附的影响,探讨其在口腔中防龋的作用。方法采用96孔微量板液体稀释法进行抑菌试验,并测得最低抑菌浓度(MIC)。体外建立变形链球菌生物膜模型,通过激光共聚焦显微镜(CLSM)扫描生物膜,观察不同浓度药物作用24 h后对生物膜活性的影响。其次测定处理后各组培养基上清液的终末pH值。最后通过玻璃棒粘附试验计算出不同浓度药物作用48 h后对生物膜粘附的影响。结果椰子油对变形链球菌的生长有抑制作用,其对变形链球菌的MIC为3.13%。CLSM观察24 h后生物膜内活菌比例逐渐下降,死菌逐渐增多。培养基上清液的终末pH值随椰子油浓度的增大而升高,且均高于阴性对照组,差异具有统计学意义(P0.05)。实验组变形链球菌的粘附率随椰子油浓度增高而降低,与阴性对照组相比差异有统计学意义(P0.05)。结论椰子油对变形链球菌有抑制作用,并能抑制其生物膜的活性、产酸及粘附等作用。     

蜂胶对变形链球菌抑菌活性的研究进展

龋病是一种微生物感染性疾病,变形链球菌是引起其发生发展的主要致龋菌之一。近年来天然药物对龋病防治的研究已成为热点,而蜂胶是一种天然抗菌剂,国内外相关研究表明蜂胶对变形链球菌的生长、产酸、粘附、产胞外多糖及牙菌斑等方面有抑制的作用。本研究就蜂胶对变形链球菌的主要致龋毒力因子的作用研究作一综述。     

大蒜素抗变异链球菌的致龋效果

目的研究大蒜素对口腔变异链球菌生长及其菌斑生物膜粘附的抑制作用。方法二倍稀释法梯度稀释测最小抑菌浓度(minimum inhibitory concentration,MIC),将MIC以上2个梯度浓度对应的培养物涂布于BHI培养基上进行次代培养获得最低杀菌浓度(minimum bactericidal concentration,MBC);酶标仪测A值观察不同浓度大蒜素抑菌效应;抑制产酸试验观察抑制细菌产酸效应;结晶紫法研究亚抑菌浓度提取物对变异链球菌粘附能力及生物膜总量的影响;采用激光共聚焦荧光显微镜(laser scanning confocal microscopy,LSCM)观察常态牙菌斑生物膜生长过程中及药物处理后牙菌斑生物膜中死菌和活菌的构成,研究其对牙菌斑生物膜结构和活性的影响。结果抑菌试验中,得到大蒜素MIC为12.8 mg/L,MBC为25.8 mg/L。MIC及亚抑菌浓度抑菌试验显示均有一定的抑菌性,抑制率为2.17%～67.12%,并且抑菌性与浓度梯度成正相关。产酸试验显示24 h内大蒜素明显抑制细菌产酸(P0.01),细菌粘附试验结果显示大蒜素在MIC时生物膜的生成速度最慢,生物膜的总量最低(P0.01)。共聚焦荧光显微镜可见大蒜素组随药物浓度增加,菌斑生物膜较薄,绿色的活菌及团块明显减少,抑制生物膜的生长。结论大蒜素对变异链球菌生长、产酸与粘附有一定抑制作用。     

天然药物对血链球菌生长和产酸影响的体外研究

目的：通过研究不同天然药物对血链球菌生长及产酸的影响,为今后筛选出能有效调理口腔菌群生态平衡的药物奠定基础。方法：选用变形链球菌SB179作为实验菌株,测定川芎,血藤,五倍子等11种天然药物的最低抑菌浓度MIC,再以低于MIC的4个浓度配制含药的TPY液体培养基,调定其初始pH为7．4,接种血链球菌,厌氧培养后测定其终末pH,结果：当药物浓度低于或等于8．000mg/ml时,各天然药物对血链球菌的生长均有一定的抑制作用,且以五倍子作用较强,槟榔,茶多酚,蜂房,大黄,三七,血藤,白芷对血链球菌的产酸具有一定的抑制能力,而黄芩,川芎,五倍子和儿茶没有明显的抑制作用,结论：槟榔,茶多酚,蜂房,大黄,三七,血藤和白芷对血链球菌的生长和产酸都有一定的抑制作用.     

黄连素对变异链球菌抑制作用的体外研究

目的分析黄连素对体外变异链球菌生长、产酸、粘附的影响,探讨其防龋作用。方法采用微量肉汤稀释法进行最小抑菌浓度(MIC)的测定;然后通过试管粘附法测定不同浓度药液对变异链球菌粘附作用的影响;最后计算不同浓度药液作用24h后pH值的变化。结果黄连素对变异链球菌的MIC为1.25mg/mL,MBC为5.00mg/mL。实验组对变异链球菌的粘附及产酸的抑制作用随着药物浓度的增加而增强,与阴性对照组相比差异有统计学意义(P0.05)。结论黄连素可抑制体外变异链球菌的生长、产酸及粘附。     

天然药物对唾液链球菌生长与产酸影响的体外研究

目的：通过研究不同天然药物对唾液链球菌生长及产酸的影响,为今后筛选出能有效调理口腔菌群生态平衡的药物奠定基础,方法：选用唾液链球菌SS196作为实验菌株,测定川芎,血藤,五倍子等11种天然药物的最低抑菌浓度MIC,再以低于MIC的4个浓度梯度配制含药的TPY液体培养基,调定其初始pH为7．4,接种唾液链球菌,厌氧培养后测定其终末pH,结果：当药物浓度低于或等于8．00mg/ml时,除槟榔,蜂房,三七外,其他药物对唾液链球菌的生长都有一定的抑制作用,且以大黄,五倍子和黄芩较强,槟榔,茶多酚,大黄,蜂房,黄芩,三七,五倍子和儿茶对唾液链球菌的产酸具有一定的抑制能力,而白芷,川芎和血藤没有明显的抑制能力,结论：茶多酚,大黄,黄芩,五倍子和儿茶对唾液链球菌的生长和产酸都有一定的抑制作用。     

茶多酚-柠檬提取物混合液对变形链球菌生长与黏附影响的研究

目的探讨茶多酚与柠檬提取物联合应用对变形链球菌生长、黏附的影响。方法采用MTT法分别测定茶多酚、柠檬提取物以及两者混合液的最小抑菌浓度(MIC)值,通过分级抑制浓度(FIC)指数来判断两者联合使用对变形链球菌生长作用的影响,并通过扫描电镜观察低于MIC浓度的混合液对变形链球菌在玻片上黏附的情况。结果茶多酚与柠檬提取物单独用药时,其MIC分别为1.56mg/mL和5.60mg/mL。联用的MIC为茶多酚0.78mg/mL和柠檬提取物4.50mg/mL,FIC为0.75。混合液对变形链球菌的黏附抑制作用随着浓度的降低而明显增强。结论茶多酚与柠檬提取物联合应用对变形链球菌的体外抗菌活性具有相加作用,影响其在玻片表面的黏附。     

赤藓糖醇对主要致龋链球菌及耐氟菌株生长和产酸的影响

目的通过赤藓糖醇对变形链球菌、远缘链球菌及其耐氟菌株混合菌生长和产酸影响的体外研究,为赤藓糖醇防龋作用的机理提供制论依据。方法采用最小抑菌浓度递增法对变形链球菌（S.mutans ATCC 25175,S.m）、远缘链球菌（S.sobrinus 6715,S.s）进行氟化钠体外诱导耐氟菌株（S.m-FR、S.s-FR）,利用液体稀释法配制赤藓糖醇TSB液8个浓度,分别加入含有变形链球菌、远缘链球菌及其耐氟菌株的细菌混悬液48 h,用比浊法观察其对混合菌生长的影响,并用pH计测定培养前后上清液的△pH值。结果吸光度A值和△pH值实验前后与对照组相比最低浓度为12%时差异均有统计学意义（P〈0.05）,且随着浓度的升高A值和△pH值均下降。结论赤藓糖醇能抑制变形链球菌、远缘链球菌及耐氟菌株混合菌生长和产酸,并且随着浓度的升高抑制作用增强。     

伊犁黑蜂蜂胶对不同状态下变形链球菌乳酸脱氢酶及其相关基因影响的实验研究

目的目的通过新疆伊犁黑蜂蜂胶乙醇提取物(Ethanol Extract of Propolis,EEP)对不同状态下变形链球菌乳酸脱氢酶活性及其相关基因表达影响的作用,研究伊犁黑蜂蜂胶抑制变形链球菌产酸的原因并探讨其可能的防龋机制。方法 (1)分别培养浮游状态与生物膜状态下生长的变形链球菌,根据实验分组用含梯度浓度EEP的BHI培养基、50 mg/L氟化钠的BHI培养基作用18 h,通过还原性辅酶I氧化法测定乳酸脱氢酶活性。(2)分别培养浮游状态与生物膜状态下生长变形链球菌,根据实验分组用含梯度浓度EEP的BHI培养基、含50 mg/L氟化钠的BHI培养基作用18 h,反转录-实时荧光定量PCR(RTq PCR)法测定各组乳酸脱氢酶编码基因ldh表达情况。结果 (1)在浮游状态与生物膜状态下,EEP组和Na F组乳酸脱氢酶活性均有降低,差异具有统计学意义(P0.05)。(2)浮游状态时,实验组组和阳性对照组ldh表达明显受到抑制(P0.05);生物膜状态下,实验组在1 MBEC、1/2 MBEC、1/4 MBEC浓度时ldh表达受到抑制(P0.05),Na F组ldh表达差异没有统计学意义(P0.05)。结论伊犁黑蜂蜂胶能够抑制浮游状态与生物膜状态下变形链球菌乳酸脱氢酶活性及其编码基因ldh表达,来抑制细菌产酸,伊犁黑蜂蜂胶可能是通过此途径抑制变形链球菌产酸,从而达到防龋的效果。     

天然植物黄蜀葵提取物对变形链球菌生长及黏附抑制作用的实验研究

目的通过研究黄蜀葵花总黄酮对变形链球菌生长及黏附的影响,为临床龋病预防提供实验基础。方法采用二倍稀释法观察不同浓度的黄蜀葵花总黄酮对变形链球菌生长的影响,测定该药物的最小抑菌浓度（MIC）;以低于MIC的5个浓度梯度配置含药的TPY液体培养基,接种变形链球菌,厌氧培养24h,计算黄蜀葵花总黄酮对变形链球菌的黏附抑制率。结果一定浓度的黄蜀葵花总黄酮能够抑制变形链球菌的生长,MIC为2．5g／L;变形链球菌的黏附率随培养基中黄蜀葵花总黄酮浓度的升高而下降,且抑菌作用呈现明显的浓度依赖性。结论天然植物黄蜀葵提取物黄蜀葵花总黄酮对变形链球菌的生长和黏附都有一定的抑制作用。     

复方茶多酚含漱液对正畸儿童牙面菌斑中致龋菌的影响

目的观察复方茶多酚含漱液对正畸儿童牙面菌斑中细菌总数和变形链球菌数的影响,以及牙菌斑内原位pH的改变。方法选择42例戴用固定矫治器的正畸儿童,随机分为2组,试验组用复方茶多酚含漱液漱口,对照组用蒸馏水漱口。分别于戴用矫治器前,戴入后1月采集上下颌牙唇颊面菌斑,测定菌斑中细菌总数及变形链球菌数,同时测定牙菌斑原位pH。结果对照组戴用后1月,细菌总数及变形链球菌数较戴用前明显增加(P0.01),牙菌斑原位pH较戴用前降低(P0.01)。试验组与对照组戴用后1个月相比,试验组细菌总数及变形链球菌数明显少于对照组(P0.01),牙菌斑原位pH高于对照组(P0.01)。结论戴用固定矫治器后,牙面菌斑内细菌总数及变形链球菌数较戴用前增加,牙菌斑原位pH较戴用前降低,应用茶多酚含漱液可明显抑制正畸儿童口腔内变形链球菌数,减少龋坏发生。     

天然药物蜂房化学成分提取物对口腔细菌生长的实验研究

目的研究蜂房中分离得到的不同组分对致龋菌生长的影响,寻找蜂房抑龋的有效成分。方法通过溶剂分段和层析技术对蜂房进行分离,得到4个组分,采用液体稀释法研究蜂房不同组分对口腔常居细菌——血液链球菌、唾液链球菌,以及4种主要致龋菌——变形链球菌、内氏放线菌、粘性放线菌和乳酸杆菌生长的影响,使用活菌计数法测量五倍子总鞣质及各组分对变形链球菌生长曲线的影响。结果蜂房提取物中1、2和3组分对实验菌有较强的抑菌作用,蜂房提取物3组分对于变形链球菌生长曲线的抑制作用最强。结论蜂房各组分对实验菌都有一定的抑菌作用,其抑菌作用可能和其中的甾醇类化合物有关。     

Molecular analysis of the inhibitory effects of oolong tea polyphenols on glucan-binding domain of recombinant glucosyltransferases from Streptococcus mutans MT8148

An oolong tea polyphenol (OTF6) has been shown to possess a strong anti-glucosyltransferase (GTF) activity and inhibit experimental dental caries in rats infected with mutans streptococci. The effects of OTF6 on the functional domains of GTFs of Streptococcus mutans, an N-terminal catalytic domain (CAT), and a C-terminal glucan-binding domain (GBD), were examined. The maximum velocity of glucan synthesis by recombinant GTFB (rGTFB) and GTFD (rGTFD) became significantly slower in the presence of OTF6, however, Km values remained stable when compared in their absence. These results suggest that OTF6 reduces glucan synthesis by non-competitively inhibiting the GBD of S. mutans GTFB and GTFD. Further, the recombinant proteins of CAT (rCAT) and GBD (rGBD) were expressed using Escherichia coli, and purified by affinity column chromatography. rGBD but not rCAT was found to possess dextran-binding activity, which was shown to be inhibited by OTF6. These results indicate that OTF6, a polymeric polyphenol specific for oolong tea is able to reduce glucan synthesis by inhibiting the GBD of S. mutans GTFB.     

Effects of compounds found in Nidus Vespae on the growth and cariogenic virulence factors of Streptococcus mutans

Nidus Vespae (honeycomb) is a kind of traditional Chinese medicine that has been demonstrated to inhibit the growth and acid-production of oral cariogenic bacteria. Subsequent studies showed that the chloroform/methanol (Chl/MeOH) chemical extraction of Nidus Vespae was the most effective inhibitor of growth and acidogenicity of Streptococcus mutans. In this study, we isolated the chemical compounds of the Nidus Vespae Chl/MeOH extraction, tested their antimicrobial activity against six cariogenic bacteria and further evaluated the acid inhibition properties, anti-F-ATPase activity and anti-LDH activity against S. mutans. The isolated flavonoids, quercetin and kaempferol, inhibited the growth of bacteria (S. mutans, Streptococcus sobrinus, Streptococcus sanguis, Actinomyces viscosus, Actinomyces naeslundii and Lactobacillus rhamnosus) with minimum inhibitory concentrations (MICs) ranging from 1 to 4 mg/ml and minimum bactericidal concentrations (MBCs) from 4 to 16 mg/ml. In addition, quercetin and kaempferol at sub-MIC levels significantly inhibited acidogenicity and acidurity of S. mutans cells. Treated with the test agents, the F-ATPase activity was reduced by 47.37% with 1mg/ml quercetin and by 49.66% with 0.5mg/ml kaempferol. The results showed that quercetin and kaempferol contained in Chl/MeOH extraction presented remarkably biological activity, suggesting that Nidus Vespae might be useful as a potential preventive and therapeutic agent in dental caries.     

Genetic exchange between oral streptococci during mixed growth

To determine whether oral streptococci might exchange genetic information in the oral cavity, paired transformable strains of Streptococcus mutans, Streptococcus sanguis and Streptococcus milleri were growth together. Chromosomal and plasmid-borne antibiotic resistance markers could be readily transferred from S. mutans GS-5 to S. milleri NCTC 10707 or S. sanguis Challis during mixed growth. However, no exchange from the latter two organisms to strain GS-5 could be detected under these conditions. The transfer of genetic information from S. sanguis to S. milleri was also observed.     

Anticariogenic activity of macelignan isolated from Myristica fragrans (nutmeg) against Streptococcus mutans

The occurrence of dental caries is mainly associated with oral pathogens, especially cariogenic Streptococcus mutans. Preliminary antibacterial screening revealed that the extract of Myristica fragrans, widely cultivated for the spice and flavor of foods, possessed strong inhibitory activity against S. mutans. The anticariogenic compound was successfully isolated from the methanol extract of M. fragrans by repeated silica gel chromatography, and its structure was identified as macelignan by instrumental analysis using 1D-NMR, 2D-NMR and EI-MS. The minimum inhibitory concentration (MIC) of macelignan against S. mutans was 3.9 microg/ml, which was much lower than those of other natural anticariogenic agents such as 15.6 microg/ml of sanguinarine, 250 microg/ml of eucalyptol, 500 microg/ml of menthol and thymol, and 1000 microg/ml of methyl salicylate. Macelignan also possessed preferential activity against other oral microorganisms such as Streptococcus sobrinus, Streptococcus salivarius, Streptococcus sanguis, Lactobacillus acidophilus and Lactobacillus casei in the MIC range of 2-31.3 microg/ml. In particular, the bactericidal test showed that macelignan, at a concentration of 20 microg/ml, completely inactivated S. mutans in 1 min. The specific activity and fast-effectiveness of macelignan against oral bacteria strongly suggest that it could be employed as a natural antibacterial agent in functional foods or oral care products.     

Role of Chlorogenic Acid Quinone and Interaction of Chlorogenic Acid Quinone and Catechins in the Enzymatic Browning of Apple

Chlorogenic acid (CQA) is one of the major polyphenols in apple and a good substrate for the polyphenol oxidase (PPO) in apple. Apple contains catechins as well as CQA, and the role of CQA quinone and its interaction with catechins in the enzymatic browning of apple were examined. Browning was repressed and 2-cysteinyl-CQA was formed when cysteine was added to apple juice. CQA quinone was essential for browning to occur. Although catechins and CQA were oxidized by PPO, some catechins seemed to be non-enzymatically oxidized by CQA quinone.