中国药物警戒 ›› 2026, Vol. 23 ›› Issue (9): 1005-1010.
DOI: 10.19803/j.1672-8629.20260434

• 基础与临床研究 • 上一篇    下一篇

共价结合蛋白化学法对5种外用原料药皮肤致敏性研究

张艳美1, 王美艳2, 阮健2, 祝清芬1,*   

  1. 1山东省食品药品检验研究院,产业技术基础公共服务平台,山东 济南 250101;
    2烟台市食品药品检验检测中心,山东 烟台 264000
  • 收稿日期:2026-06-02 出版日期:2026-09-15 发布日期:2026-09-15
  • 通讯作者: *祝清芬,男,博士,主任药师,药品检验。E-mail: zhuqingfen73@163.com
  • 作者简介:张艳美,女,硕士,高级工程师,药品检验。
  • 基金资助:
    国家重点研发计划（2022YFF0711105）; 山东省药品监督管理局检验检测能力提升项目（SDNMPAFZLY202501）

Skin sensitization of five topical active pharmaceutical ingredients based on a chemical method for covalently binding proteins

Zhang Yanmei1, Wang Meiyan2, Ruan Jian2, Zhu Qingfen1,*   

  1. 1Industrial Technology Foundation Public Service Platform, Shandong Institute for Food and Drug Control, Jinan Shandong 250101, China;
    2Yantai Food and Drug Inspection and Testing Center, Yantai Shandong 264000, China
  • Received:2026-06-02 Online:2026-09-15 Published:2026-09-15

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摘要： 目的 建立共价结合蛋白化学法对5种外用化学原料进行皮肤致敏性评价。方法 用直接多肽反应试验（DPRA）和动力学直接多肽反应试验（kDPRA）对常用的5种外用化学原料药（米诺地尔、洛索洛芬钠、丙胺卡因、利多卡因、氟比洛芬）进行皮肤致敏性检测。参照经济合作与发展组织（OECD）TG442C选择2种物质验证DPRA和kDPRA;将5种原料药分别与半胱氨酸多肽和赖氨酸多肽模拟的皮肤蛋白共同孵育,运用高效液相色谱（HPLC）法测定多肽含量,根据多肽消耗百分比评估其皮肤致敏性;筛选出3种原料药（与赖氨酸多肽发生共洗脱）仅与半胱氨酸多肽溶液孵育不同时间后,运用酶标仪测定荧光强度,根据反应动力学常数对其进行致敏性分级。结果 实验室验证合格率100%,表明实验系统稳定可靠。洛索洛芬钠、氟比洛芬多肽消耗率均小于6.38%,致敏性为阴性。利多卡因、丙胺卡因显示非反应性,致敏性为阴性;米诺地尔lgkmax< -2.0,为中、弱致敏剂。结论 DPRA和kDPRA相结合的化学法,方法互补,操作简单,可作为外用化学原料药皮肤致敏性初步评价的新方法。

关键词: 米诺地尔, 洛索洛芬钠, 丙胺卡因, 利多卡因, 氟比洛芬, 外用化学原料药, 皮肤致敏性, 直接多肽反应试验, 动力学直接多肽反应试验

Abstract: Objective To establish a chemical method for covalently binding proteins and evaluate the skin sensitization of five topical chemical materials. Methods The chemical method using covalent binding proteins (direct peptide reaction test or kinetic direct peptide reaction test) was employed to conduct skin sensitization tests on five commonly used topical chemical raw materials including minoxidil, loxoprofen sodium, prilocaine, lidocaine and flurbiprofen in pharmaceuticals. According to OECD TG442C, two substances were selected to validate the direct peptide reaction test (DPRA) and kinetic direct peptide reaction test (kDPRA) in our laboratory. These five raw materials were incubated with cysteine peptide and lysine peptide mimicking skin proteins respectively, and the peptide content was determined using high-performance liquid chromatography. Skin sensitization was evaluated based on the percentage of peptide consumption. Three of these raw materials (co-eluted with lysine peptide) were incubated with the cysteine peptide solution for different lengths of time before the fluorescence intensity was measured using an enzyme-labeled instrument to classify their sensitization based on the reaction kinetic constant. Results The pass rate was 100% based on laboratory verification, suggesting that the experimental system was stable and reliable. The consumption rates of loxoprofen sodium and flurbiprofen peptide were both below 6.38%, while sensitization was negative. Lidocaine and piperacaine were non-reactive and negative in sensitization, compared with lgkmax <-2.0 for minoxidil, indicating it was a moderate or weak sensitizing agent. Conclusion The combined chemical method of DPRA and kDPRA is complementary and user-friendly. It can be used as a new method for preliminary evaluation of skin sensitization of topical chemical raw materials.

Key words: Minoxidil, Loxoprofen Sodium, Prilocaine, Lidocaine, Flurbiprofen, Topical Chemical Active Pharmaceutical Ingredients, Skin Sensitization, DPRA, kDPRA

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