Study of Antibacterial Substances Found in Natto

采用Janssen等人的比色法对枯草芽孢杆菌纳豆细胞中二吡啶酸（DPA,2,6-吡啶二羧酸）含量进行分析，DPA含量最高，约占总干重的2.4-3.6%。对DPA及其衍生物喹啉酸（2,3-吡啶二羧酸）、2,4-吡啶二羧酸、异绿酸（2,5-吡啶二羧酸）和3,5-吡啶二羧酸的抑菌活性进行了比较，发现DPA具有独特的抑菌活性。此外，研究了各种金属离子对抗菌活性的影响，结果表明，Ca^<++>、Fe^<++>、Co^<++>和Zn^<++>均为二价离子，抗菌活性被强烈抑制。实验还表明，添加DPA可提高枯草芽孢杆菌纳豆产生纳豆激酶的量。在宫野型、黑黑型、高桥型和成濑型4种枯草芽孢杆菌纳豆中，DPA浓度为2 mM时，纳豆激酶的产生量增加。例如，在宫野型的情况下，纤维蛋白板的纤溶活性是未添加DPA的样品的两倍，在37℃下，4小时后达到256.1 mm^2/30 μl（纳豆激酶活性为1,500 FU/ml）。通过添加10种烟酸相关物质的对比试验，发现二吡啶酸的效果最强。发现终浓度5×10^<-3> mol/l的DPA能显著抑制人三磷酸腺苷（ATP）引起的血小板聚集反应，以及凝血酶将纤维蛋白原转化为纤维蛋白的反应。很明显，在5×10^<-3> mol/l时，血栓弹性成像中观察到的凝血反应完全被抑制，而DPA衍生物即使在10 mM时也没有观察到这种强烈的作用。人凝血酶原时间、活性区域的凝血活素时间或优球蛋白溶解时间没有变化。虽然在30 μM的二磷酸腺苷（ADP）下观察到人血液中的血小板聚集，但很明显，当DPA浓度为5×10^<-3> M时，这种聚集发生剂量依赖性抑制。计算出的CI_<50>浓度为1.8×10^<-3> M，较低，远高于加入阿司匹林临床使用时的浓度（4.0×10^<-2> M）。少

An analysis of dipicolinic acid (DPA,2,6-pyridinedicarboxylic acid) contained in Bacillus subtilis natto cells using the colorimetric method of Janssen and others showed that the largest DPA content is approximately 2.4-3.6% of the total dry weight.A comparison of the antibacterial activity of DPA and its derivatives-quinolic acid (2,3-pyridine dicarboxylic acid), 2,4-pyridine dicarboxylic acid, isoinchlorendic acid (2,5-pyridine dicarboxylic acid), and 3,5-pyridine dicarboxylic acid-revealed that this antibacterial activity is unique to DPA. Moreover, an investigation into the effects of various metal ions on the antibacterial activity showed that the activity is strongly inhibited through the addition of Ca^<++>,Fe^<++>,Co^<++>, and Zn^<++>, all divalent ions.It was also shown that the addition of DPA increases the amount of nattokinase produced by Bacillus subtilis natto. In four types of Bacillus subtilis natto, the Miyagino, Meguro, Takahashi and Naruse types, DPA with a 2 mM conc … More entration increased the amount of nattokinase produced. In the case of the Miyagino type, for example, the fibrinolytic activity for fibrin plates was double the level seen for the sample with no DPA added, at 256.1 mm^2/30 μl at 37℃ after four hours (1,500 FU/ml for the nattokinase activity). A comparison test, with 10 types of nicotinic acid-related substances added, showed that dipicolinic acid had the strongest effect.DPA with a final concentration of 5×10^<-3> mol/l was found to substantially inhibit the platelet aggregation reaction caused by human adenosine triphosphate(ATP), as well as the reaction of thrombin converting fibrinogen into fibrin. It became clear that at 5×10^<-3> mol/l, there was a complete inhibition of the blood coagulation reaction as observed in thromboelastography, while such strong effects were not observed in the case of DPA derivatives, even at 10 mM. There were no changes in human prothrombin time, the thromboplastin time for the active areas, or euglobulin lysis time.Although platelet aggregation in human blood is observed with 30 μM of adenosine diphosphate(ADP), it became clear that dose-dependent inhibition of this aggregation occurs when there is 5×10^<-3> M of DPA. The calculated CI_<50> was at the fairly low concentration of 1.8×10^<-3> M, and far stronger (4.0×10^<-2> M) than in the case of adding aspirin for clinical use. Less