Department of Pharmacognosy, Faculty of Pharmacy with the Division of Laboratory Medicine, Medical University of Białystok, ul. Mickiewicza 2a, 15-230 Białystok, Poland.
Department of Synthesis and Technology of Drugs, Faculty of Pharmacy with the Division of Laboratory Medicine, Medical University of Białystok, ul. Kilińskiego 1, 15-089 Białystok, Poland.
Int J Mol Sci. 2021 Mar 25;22(7):3345. doi: 10.3390/ijms22073345.
L. (Campanulaceae) is used in traditional Belarusian herbal medicine for sleep disorders in children, but the chemical composition and biological activity have not been investigated. In this study, the activities of extracts, their fractions and main compounds were evaluated in amelanotic melanoma C32 (CRL-1585) cells and normal fibroblasts (PCS-201-012). The extracts and fractions were analyzed using liquid chromatography-photodiode array detection-electrospray ionization-mass spectrometry (LC-PDA-ESI-MS/TOF) to characterize 25 compounds. Further, three major and known constituents, luteolin () and its derivatives such as 7--glucoside () and 7--sambubioside () were isolated and identified. The cytotoxic activities against fibroblasts and the amelanotic melanoma cell line were determined using the fixable viability stain (FVS) assay. The influence of diethyl ether (EtO) fraction () and on apoptosis induction was investigated using an annexin V binding assay. The obtained results showed significant cytotoxicity of and with IC values of 119.7 ± 3.2 and 95.1 ± 7.2 μg/mL, respectively. The proapoptotic potential after treatment in the C32 human amelanotic melanoma cell line was comparable to that of vinblastine sulfate (VLB), detecting 29.2 ± 3.0% apoptotic cells. Moreover, displayed less necrotic potential against melanoma cells than VLB. In addition, the influences of and on the dysfunction of the mitochondrial membrane potential (MMP), cell cycle and activity of caspases 3, 8, 9, and 10 were established. The effects of on MMP change (74.5 ± 3.0% of the cells showed a reduced MMP) corresponded to the results obtained from the annexin V binding assay and activation of caspase-9. and displayed a significant impact on caspase-9 (40.9 ± 2.4% of the cells contained active caspase-9 after treatment and 16.6 ± 0.8% after incubation with ) and the intrinsic (mitochondrial) apoptotic pathway. Moreover, studies have shown that and affect the activation of external apoptosis pathways by inducing the caspase-8 and caspase-10 cascades. Thus, activation of caspase-3 and DNA damage via external and internal apoptotic pathways were observed after treatment with and . The obtained results suggest that extracts could be developed as new topical preparations with potential anticancer properties due to their promising cytotoxic and proapoptotic potential.
（桔梗科）在传统的白俄罗斯草药医学中用于治疗儿童睡眠障碍，但尚未对其化学成分和生物活性进行研究。在这项研究中，评估了提取物、其馏分和主要化合物在无黑色素性黑色素瘤 C32（CRL-1585）细胞和正常成纤维细胞（PCS-201-012）中的活性。使用液相色谱-光电二极管阵列检测-电喷雾电离质谱（LC-PDA-ESI-MS/TOF）分析提取物和馏分，以鉴定 25 种化合物。此外，分离并鉴定了三种主要的已知成分，木犀草素（）及其衍生物如 7--葡萄糖苷（）和 7--桑布糖苷（）。使用固定化生存力染色（FVS）测定法测定了对成纤维细胞和无黑色素性黑色素瘤细胞系的细胞毒性。使用 Annexin V 结合测定法研究了二乙醚（EtO）馏分（）和 对细胞凋亡诱导的影响。结果表明，和 具有显著的细胞毒性，IC 值分别为 119.7±3.2 和 95.1±7.2μg/mL。在 C32 人无黑色素性黑色素瘤细胞系中，处理后的促凋亡潜力与硫酸长春碱（VLB）相当，检测到 29.2±3.0%的凋亡细胞。此外，与 VLB 相比，对黑素瘤细胞的坏死潜力较小。此外，还确定了和 对线粒体膜电位（MMP）、细胞周期和半胱天冬酶 3、8、9 和 10 活性的影响。（74.5±3.0%的细胞显示 MMP 降低）与 Annexin V 结合测定和 caspase-9 激活的结果相对应。和 对 caspase-9 （处理后 40.9±2.4%的细胞含有活性 caspase-9，用处理后 16.6±0.8%）和内在（线粒体）凋亡途径有显著影响。此外，研究表明和 通过诱导 caspase-8 和 caspase-10 级联反应，影响外源性凋亡途径的激活。因此，在用和 处理后，通过外源性和内源性凋亡途径观察到 caspase-3 和 DNA 损伤的激活。研究结果表明，由于其有希望的细胞毒性和促凋亡潜力，提取物可以开发为具有潜在抗癌特性的新型局部制剂。
