Development and evaluation of berberine-loaded bigel for the treatment of hyperpigmentation on B16F10 melanoma cell line

<b>Aim:</b> The aim of this study was to optimize, develop, characterize and evaluate a topical nanobigel (BG) formulation containing Berberine (BRB) that exhibits anti-melanogenic properties. <b>Materials &amp; methods:</b> The Berberine-loaded bigel (BRB@BG) formulation was prepared by homogenously mixing the optimized hydrogel and oleogel. BRB@BG was characterized <i>in vitro</i> and cytotoxicity study was conducted to evaluate its effects on murine skin melanoma B16F10 cell lines. <b>Results:</b> The optimized BRB@BG exhibited uniform texture with nanometric size, desirable spreadability and extrudability, suitable for topical applications. Cytotoxicity studies revealed that BRB@BG had a lower IC₅₀ value (4.84 μg/ml) on B16F10 cell lines compared with drug alone. <b>Conclusion:</b> In conclusion, the developed BRB@BG formulation showed good potential as safe and effective topical treatment for hyperpigmentation. Hyperpigmentation is a common skin disease in which the melanin content becomes abnormally high. The existing treatment options are associated with side effects and therefore research is being aimed to develop a topical nanoformulation based on natural compounds. Berberine (BRB) is one such natural compound that is known to inhibit the production of melanin in skin. This study was aimed to develop a bigel formulation of BRB that can be applied on skin, which can treat the hyperpigmentation and is safe. The developed nanomedicine was found to have all good properties of a topical formulation and was more effective than the drug alone. Various studies on animals were conducted to assess its safety and it was found that the formulation did not show any toxicity to the skin. Berberine, an isoquinoline alkaloid is reported to have antimelanogenic effect but is associated with poor oral bioavailability. Berberine-loaded bigel (BRB@BG) was prepared by homogenously mixing the optimized hydrogel and oleogel. The prepared BRB@BG was characterized using <i>in vitro</i> methods, TEM and trinocular microscopy. Cytotoxicity study was conducted to evaluate the effects of BRB@BG on the murine skin melanoma B16F10 cell lines. Safety of the developed formulation was performed using Draize test followed by histopathology and <i>in ovo</i> irritation studies. The developed formulation showed all desirable properties of an ideal topical formulation. The IC₅₀ values obtained after cytotoxicity studies was found to be lower than the plain drug showing better efficacy. The developed formulation showed no toxicity in any of the safety evaluation test.

**研究目的**：本研究旨在优化、开发、表征并评价一款含小檗碱（Berberine, BRB）的局部外用纳米双凝胶（nanobigel, BG）制剂，该制剂具备抗黑素生成特性。
**材料与方法**：通过将优化后的水凝胶与油凝胶均匀混合，制备载小檗碱双凝胶（BRB@BG）。采用体外（in vitro）实验对BRB@BG进行表征，并以小鼠皮肤黑色素瘤B16F10细胞系为模型开展细胞毒性实验，评估其生物学效应。
**结果**：优化后的BRB@BG质地均匀，粒径呈纳米级，具备良好的铺展性与可挤出性，适用于局部外用。细胞毒性实验结果显示，相较于游离小檗碱，BRB@BG对B16F10细胞的半数抑制浓度（IC₅₀）为4.84 μg/ml，显著更低。
**结论**：本研究开发的BRB@BG制剂具备安全有效的局部治疗色素沉着过度的潜力。
色素沉着过度是一类常见皮肤疾病，患者皮肤黑色素含量异常升高。现有治疗手段多伴随不良反应，因此基于天然化合物开发外用纳米制剂的研究备受关注。小檗碱是此类天然化合物之一，其可抑制皮肤黑色素生成。本研究旨在开发可用于皮肤的小檗碱双凝胶制剂，以治疗色素沉着过度且具备安全性。所开发的纳米制剂具备外用制剂的各项优良特性，疗效优于游离小檗碱。多项动物实验用于评估其安全性，结果显示该制剂无皮肤毒性。小檗碱作为异喹啉类生物碱，已被报道具备抗黑素生成活性，但口服生物利用度较差。本研究所制备的BRB@BG通过体外实验、透射电子显微镜（TEM）及三目显微镜完成表征，细胞毒性实验用于评估其对B16F10细胞的效应，安全性评价则通过德里兹皮肤刺激性试验（Draize test）、组织病理学检查及鸡胚内（in ovo）刺激性试验完成。所开发的制剂具备理想外用制剂的各项优良特性，细胞毒性实验所得半数抑制浓度低于游离药物，显示出更优的疗效，且所有安全性评价试验均未发现其存在毒性。