Sawant Sheeba, Baldwin Timothy C, Metryka Oliwia, Rahman Ayesha
Faculty of Science and Engineering, University of Wolverhampton, Wulfruna St., Wolverhampton WV1 1LY, UK.
Doctoral School, University of Silesia, Bankowa 14, 40-032 Katowice, Poland.
Pathogens. 2023 Jun 20;12(6):853. doi: 10.3390/pathogens12060853.
(Indian borage) has been extensively studied for its medicinal properties, which can be exploited to develop new antimicrobial therapeutics. The current study investigated the effect of leaf extracts on the catalase activity, reactive oxygen species, lipid peroxidation, cytoplasmic membrane permeability, and efflux pump activity in NCTC8325 and PA01. As the enzyme catalase protects bacteria against oxidative stress, disruption of its activity creates an imbalance in reactive oxygen species (ROS) levels, which subsequently oxidizes lipid chains, leading to lipid peroxidation. In addition, bacterial cell membranes are a potential target for new antibacterial agents, as efflux pump systems play a crucial role in antimicrobial resistance. Upon exposure of the microorganisms to Indian borage leaf extracts, the observed catalase activity decreased by 60% and 20% in and respectively. The generation of ROS can cause oxidation reactions to occur within the polyunsaturated fatty acids of the lipid membranes and induce lipid peroxidation. To investigate these phenomena, the increase in ROS activity in and was studied using HDCFDA, which is oxidized to 2',7'-dichlorofluorescein (DCF) by ROS. Furthermore, the concentration of lipid peroxidation product (malondialdehyde) was assessed using the Thiobarbituric acid assay and was shown to increase by 42.4% and 42.5% in and respectively. The effect of the extracts on the cell membrane permeability was monitored using diSC3-5 dye and it was observed that the cell membrane permeability of increased by 58% and of by 83%. The effect on efflux pump activity was investigated using Rhodamine-6-uptake assay, which displayed a decrease in efflux activity of 25.5% in and 24.2% in after treatment with the extracts. This combination of different methods to study various bacterial virulence factors provides a more robust, mechanistic understanding of the effect of extracts on and . This study thus represents the first report of the assessment of the effect of Indian borage leaf extracts on bacterial antioxidant systems and bacterial cell membranes, and can facilitate the future development of bacterial resistance modifying agents derived from .
（印度琉璃苣）因其药用特性已得到广泛研究，这些特性可用于开发新的抗菌疗法。当前研究调查了叶提取物对NCTC8325和PA01中过氧化氢酶活性、活性氧、脂质过氧化、细胞质膜通透性及外排泵活性的影响。由于过氧化氢酶可保护细菌免受氧化应激，其活性的破坏会导致活性氧（ROS）水平失衡，进而氧化脂质链，引发脂质过氧化。此外，细菌细胞膜是新型抗菌剂的潜在靶点，因为外排泵系统在抗菌耐药性中起关键作用。将微生物暴露于印度琉璃苣叶提取物后，观察到NCTC8325和PA01中的过氧化氢酶活性分别降低了60%和20%。活性氧的产生可导致脂质膜的多不饱和脂肪酸发生氧化反应并诱导脂质过氧化。为研究这些现象，使用HDCFDA研究了NCTC8325和PA01中活性氧活性的增加，HDCFDA会被活性氧氧化为2',7'-二氯荧光素（DCF）。此外，使用硫代巴比妥酸测定法评估脂质过氧化产物（丙二醛）的浓度，结果显示NCTC832 and PA01中分别增加了42.4%和42.5%。使用diSC3-5染料监测提取物对细胞膜通透性的影响，观察到NCTC8325的细胞膜通透性增加了58%，PA01的增加了83%。使用罗丹明-6摄取测定法研究对外排泵活性的影响，结果显示提取物处理后，NCTC8325的外排活性降低了25.5%，PA01的降低了24.2%。这种结合不同方法研究各种细菌毒力因子的方式，能更全面、深入地了解提取物对NCTC8325和PA01的影响。因此，本研究首次报道了评估印度琉璃苣叶提取物对细菌抗氧化系统和细菌细胞膜的影响，有助于未来开发源自印度琉璃苣的细菌耐药性修饰剂。
