Haplophyllum tuberculatum is a plant commonly used in folk medicine to treat several diseases including vomiting, nausea, infections, rheumatism, and gastric pains. In the current study, H. tuberculatum essential oils, hydrosols, the pure compounds R-(+)-limonene, S-(−)-limonene, and 1-octanol, as well as their combinations R-(+)-limonene/1-octanol and S-(−)-limonene/1-octanol, were screened for their cytotoxicity on HEp-2 cells after 24, 48, and 72 h, and then tested for their activity against Coxsackievirus B3 and B4 (CV-B3 and CV-B4) at 3 different moments: addition of the plant compounds before, after, or together with virus inoculation. Results showed that the samples were more cytotoxic after 72 h than after 24 h or 48 h cell contact. However, the combinations R-(+)-limonene/1-octanol and S-(−)-limonene/1-octanol showed less effect on HEp-2 cells than pure R-(+)-limonene and S-(−)-limonene after 24 h, 48 h, and 72 h. 1-octanol exhibited the highest concentration causing 50% cytotoxicity (CC50) on HEp-2 cells after 24 h (CC50 = 93 µg/mL) and 48 h (CC50 = 83 µg/mL). The antiviral assays showed that the tested samples exhibited potent inhibition of CV-B. IC50 values ranged from 0.66 µg/mL to 28.4 µg/mL. In addition, CV-B3 was more sensitive than CV-B4. Both CV-B strains are more inhibited when cells were pretreated with the plant compounds. The hydrosols have no effect, neither on HEp-2 cells nor on the virus. 1-octanol, S-(−), and R-(+)-limonene/1-octanol had important selectivity indexes over time. Although essential oils had potent antiviral activity, they can be considered for application in the pretreatment of cells. However, 1-octanol and the combinations are within the safety limits, and thus, they can be used as an active natural antiviral agent for CV-B3 and CV-B4 inhibition.

中文翻译：

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Haplophyllum tuberculatum 精油、纯化合物及其组合对柯萨奇病毒 B3 和 B4 的细胞毒性和抗病毒活性

Haplophyllum tuberculatum 是一种常用于民间医学的植物，用于治疗多种疾病，包括呕吐、恶心、感染、风湿病和胃痛。在目前的研究中，结核菌精油、纯露、纯化合物 R-(+)-柠檬烯、S-(-)-柠檬烯和 1-辛醇，以及它们的组合 R-(+)-柠檬烯/在 24、48 和 72 小时后筛选 1-辛醇和 S-(-)-柠檬烯/1-辛醇对 HEp-2 细胞的细胞毒性，然后测试它们对柯萨奇病毒 B3 和 B4 (CV-B3和 CV-B4) 在 3 个不同时刻：在病毒接种之前、之后或与病毒接种一起添加植物化合物。结果表明，样品在 72 小时后比细胞接触 24 小时或 48 小时后更具细胞毒性。然而，R-(+)-柠檬烯/1-辛醇和S-(-)-柠檬烯/1-辛醇的组合对HEp-2细胞的影响小于纯R-(+)-柠檬烯和S-(-)-柠檬烯24 小时、48 小时和 72 小时后。1-辛醇在 24 小时 (CC50 = 93 µg/mL) 和 48 小时 (CC50 = 83 µg/mL) 后显示出对 HEp-2 细胞产生 50% 细胞毒性 (CC50) 的最高浓度。抗病毒试验表明，测试样品表现出对 CV-B 的有效抑制。IC50 值范围为 0.66 µg/mL 至 28.4 µg/mL。此外，CV-B3 比 CV-B4 更敏感。当用植物化合物预处理细胞时，两种 CV-B 菌株都受到更大的抑制。水溶胶对 HEp-2 细胞和病毒都没有影响。随着时间的推移，1-辛醇、S-(-) 和 R-(+)-柠檬烯/1-辛醇具有重要的选择性指标。虽然精油具有强大的抗病毒活性，可以考虑将它们用于细胞的预处理。然而，1-辛醇及其组合在安全范围内，因此，它们可以用作抑制 CV-B3 和 CV-B4 的活性天然抗病毒剂。