This study focused on the investigation of photodegradation of usnic acid (UA) which is a natural UV absorbing agent in lichens. Despite years of historical use in food supplement, traditional medicine or cosmetic products in many countries, liver toxicity has been found to be one of the severe and life threatening adverse effects in early 2000's. Such severe side effect has limited UA or its synthesized derivatives for further use clinically or commercially. In this study, extracted UA from Usnea longissima in methanol was exposed to natural sunlight for 21 days. Five photodegraded derivatives (1 to 5) with two new and three previously explored compounds were isolated and purified by column chromatography and preparative liquid chromatography. The structures of these derivatives were identified based on the data of nuclear magnetic resonance spectrum, mass spectrum, optical rotation, infrared spectrum, X-ray crystallography and/or electronic circulation dichroism. The cytotoxicity of (+)-UA and 2 to 5 in liver L02 cells and melanocytes were evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Experimental results indicated that IC50 of (+)-UA in liver L02 cells and melanocytes were 24.4 and 6.9 μM respectively, while compound 2 to 5 have lower cytotoxicity with IC50 of 326.7, 1085.0, 62.7 and 152.4 μM in L02 cells and 87.7, 297.7, 60.2 and 85.0 μM in melanocytes respectively. Besides, (+)-UA and these derivatives were exposed to fix dosed of UVA or UVB. The anti-UVA/UVB activity was determined via Hoechst33342/propidium iodide double staining method, and quantified by computer linked fluorescence microscope equipped with CellsSense Dimension system. Based on analysis, Compound 2 to 5 captured prominent UVA/UVB protection capacity in both hepatocytes and melanocytes (p < .001). In addition, the effects of chemicals on tyrosinase were evaluated via Western Blot analysis. In terms of tyrosinase expression, only 2 showed significant stimulating effect (p < .05). However, the safe use of these derivatives cutaneously should be further studied. In conclusion, the photodegraded derivatives (2 to 5) of extracted UA have lower hepatotoxicity than (+)-UA and captured significant UV protection activities.

中文翻译：

---

在正常人L02肝细胞和表皮黑素细胞中鉴定具有改进的毒性特征和UVA / UVB保护的松萝酸光降解衍生物。

这项研究的重点是研究地衣中的天然紫外线吸收剂松萝酸（UA）的光降解作用。尽管在许多国家中在食品补充剂，传统医学或化妆品中已有多年使用历史，但在2000年代初期，人们发现肝脏毒性是严重的且威胁生命的不良反应之一。这种严重的副作用限制了UA或其合成衍生物的临床或商业用途。在这项研究中，从松萝中提取的UA在甲醇中暴露于自然阳光下21天。通过柱色谱法和制备液相色谱法分离并纯化了五种具有两种新化合物和三种先前探索过的化合物的光降解衍生物（1-5）。基于核磁共振光谱，质谱，旋光性，红外光谱，X射线晶体学和/或电子循环二色性的数据来​​鉴定这些衍生物的结构。通过3-（4，5-二甲基噻唑-2-基）-2，5-二苯基溴化四唑（MTT）测定法评价（+）-UA和2至5在肝L02细胞和黑素细胞中的细胞毒性。实验结果表明（+）-UA在肝L02细胞和黑素细胞中的IC50分别为24.4和6.9μM，而化合物2至5的细胞毒性较低，在L02细胞中的IC50为326.7、1085.0、62.7和152.4μM，分别为87.7、297.7 ，分别在黑色素细胞中为60.2和85.0μM。此外，将（+）-UA和这些衍生物暴露于固定剂量的UVA或UVB中。通过Hoechst33342 /碘化丙啶双重染色法测定抗UVA / UVB活性，并通过配备CellsSense Dimension系统的计算机链接荧光显微镜进行定量。根据分析，化合物2至5在肝细胞和黑素细胞中均具有出色的UVA / UVB保护能力（p <.001）。此外，还通过Western Blot分析评估了化学品对酪氨酸酶的影响。就酪氨酸酶表达而言，只有2个显示出明显的刺激作用（p <.05）。但是，应进一步研究这些衍生物在皮肤上的安全使用。总之，提取的UA的光降解衍生物（2至5）具有比（+）-UA更低的肝毒性，并具有明显的紫外线防护活性。化合物2至5在肝细胞和黑素细胞中均具有突出的UVA / UVB保护能力（p <.001）。此外，还通过Western Blot分析评估了化学品对酪氨酸酶的影响。就酪氨酸酶表达而言，只有2个显示出明显的刺激作用（p <.05）。但是，应进一步研究这些衍生物在皮肤上的安全使用。总之，提取的UA的光降解衍生物（2至5）具有比（+）-UA更低的肝毒性，并具有明显的紫外线防护活性。化合物2至5在肝细胞和黑素细胞中均具有显着的UVA / UVB保护能力（p <.001）。此外，还通过Western Blot分析评估了化学品对酪氨酸酶的影响。就酪氨酸酶表达而言，只有2个显示出明显的刺激作用（p <.05）。但是，应进一步研究这些衍生物在皮肤上的安全使用。总之，提取的UA的光降解衍生物（2至5）具有比（+）-UA更低的肝毒性，并具有明显的紫外线防护活性。