目的:采用高效液相色谱串联飞行时间质谱法高通量筛查化妆品中的非法添加成分,并建立高效液相色谱串联三重四极杆质谱法测定化妆品中他克莫司和吡美莫司的方法。方法:通过高效液相色谱串联飞行时间高分辨质谱法对市售儿童化妆品进行风险物质高通量初步筛查,发现1批他克莫司阳性样品,然后运用高效液相色谱串联三重四极杆质谱法进行定性定量测定。经实验条件优化,以含17.5 mmol·L-1乙酸-5 mmol·L-1乙酸铵的水溶液(A)和含17.5 mmol·L-1乙酸-5 mmol·L-1乙酸铵的甲醇溶液(B)为流动相,采用Waters ACQUITY UPLC BEH C18色谱柱,进行梯度洗脱分离,柱温45 ℃,流速0.3 mL·min-1,然后经三重四极杆质谱正离子多反应监测模式扫描。结果:他克莫司和吡美莫司质量浓度在0.5~50 ng·mL-1范围内线性关系良好,检出限均为10 ng·g-1,定量限均为25 ng·g-1。在膏霜、乳液、水剂、油剂4种代表基质中各浓度水平加标回收率均在93.1%~110.7%,回收率良好。经测定,1批儿童化妆品中检出他克莫司,含量为2.8 mg·kg-1。结论:该方法弥补了化妆品中非法添加的他克莫司及吡美莫司检验方法的缺失,为化妆品的日常监督监管提供了技术支持。
Objective: To establish a method for the determination of tacrolimus and pimeclimus in cosmetics by high performance liquid chromatography with a triple quadruple tandem mass spectrometry (HPLC-MS/MS), and was used to high-throughput screening for illegally added ingredients in cosmetics by high performance liquid chromatography-time-of-flight mass spectrometry (HPLC-Q TOF MS/MS). Methods: Through high-throughput screening of the initial risk assessment of children’s cosmetics sold in markets by HPLC-Q TOF MS/MS, a batch of tacrolimus positive children’s cosmetics was identified, and then a method was established for qualitative and quantitative determination by HPLC-MS/MS. After the optimization of chromatographic analysis conditions, the gradient elution was performed on a Waters ACQUITY UPLC BEH C18 column at 45 ℃, which were used an aqueous solution (containing 17.5 mmol·L-1 acetic acid -5 mmol·L-1 ammonium acetate) and a methanol solution (containing 17.5 mmol·L-1 acetic acid-5 mmol·L-1 ammonium acetate) as mobile phases at the flow rate of 0.3 mL·min-1. The analytes were scanned by HPLC-MS/MS in positive ion multiple reaction monitoring mode. Results: There was a good linear relationship between Tacrolimus and pimecrolimus in the concentration range of 0.5-50 ng·mL-1.The limits of detection were 10 ng·g-1and the limits of quantification concentration were 25 ng·g-1. The spiked recoveries of each concentration level were good, which ranged from 93.1% to 110.7% in the four respective matrices, such as cream, emulsion, water and oil. Tacrolimus was detected in 1 batch of children’s cosmeticsat 2.8 mg·kg-1. Conclusion: This method makes up for the lack of test methods for tacrolimus and pimecrolimus illegally addition in cosmetics, and provides technical supports for the daily supervision in cosmetics.
[1] 胡珊. 特应性皮炎诊疗研究的进展[J]. 中国当代医药, 2021, 28(28):25
HU S. Advances in diagnosis and treatment of atopic dermatitis[J]. China Mod Med, 2021, 28(28):25
[2] 曹华. 他克莫司在皮肤科的临床应用[J]. 皮肤科学通报, 2021, 38(4):343
CAO H. Clinical application of tacrolimus in dermatology[J]. Dermatol Bull, 2021, 38(4):343
[3] NGHIEM P, PEARSON G, LANGLEY RG. Tacrolimus and pimecrolimus: from clever prokaryotes to inhibiting calcineurin and treating atopic dermatitis[J]. J Am Acad Dermatol, 2002, 46(2):228
[4] 国家药典委员会.中华人民共和国药典临床用药须知 化学药和生物制品卷[M]. 北京: 中国医药科技出版社, 2015
State Pharmacopoeia Commission. The Pharmacopoeia of the People’s Republic of Clinical Medication Notice Chemical Drugs and Biological Products Volume[M]. Beijing: China Medical Science Press, 2015
[5] CHIA BK, TEY HL. Systematic review on the efficacy, safety, and cost-effectiveness of topical calcineurin inhibitors in atopic dermatitis[J]. Dermatitis, 2015, 26(3):122
[6] 刘玲玲, 窦侠, 谢志强, 等. 他克莫司软膏治疗儿童特应性皮炎疗效和安全性研究[J]. 中华皮肤科杂志, 2005, 38(10):608
LIU LL, DOU X, XIE ZQ, et al. Efficacy and safety of tacrolimus ointment for the treatment of atopic dermatitis in Chinese children[J]. Chin J Dermatol, 2005, 38(10):608
[7] 汪科, 朱学骏. 局部免疫调节剂他克莫司、吡美莫司在皮肤病的应用[J]. 药物不良反应杂志, 2005, 7(5):345
WANG K, ZHU XJ. Topical immunomodulators tacrolimus and pimecrolimus used for dermatosis[J]. Adv Drug React J, 2005, 7(5):345
[8] 白文永. 他克莫司与吡美莫司治疗特应性皮炎的系统评价[D]. 济南: 山东大学, 2009
BAI WY. Systematic Review of Tacrolimus and Pimecrolimus in The Treatment of Atopic Dermatitis[D]. Jinan: Shandong University, 2009
[9] 蒋冲. 长期外用他克莫司软膏对特应性皮炎患者免疫力的影响[D]. 广州: 南方医科大学, 2013
JIANG C. The Influence of Long-Term Topical Tacrolimus Ointment on Immunity of Patients with Atopic Dermatitis[D]. Guangzhou: Southern Medical University, 2013
[10] 吴景, 邢书霞, 王钢力. 化妆品中非法添加禁用物质检测技术研究进展[J]. 日用化学品科学, 2015, 38(10):5
WU J, XING SX, WANG GL. Research progress on determination technologies for prohibited substances illegally added in cosmetics[J]. Deterg Cosmet, 2015, 38(10):5
[11] 王延东, 唐细兰, 叶成添, 等. 高效液相色谱法测定吡美莫司滴眼液的含量[J]. 中国新药杂志, 2010, 19(21):2005
WANG YD, TANG XL, YE CT, et al. Determination of pimecrolimus in eye drops by HPLC[J]. Chin New Drugs J, 2010, 19(21):2005
[12] 陈悦. HPLC法测定他克莫司的含量及有关物质[J]. 药物分析杂志, 2008, 28(2):309
CHEN Y. HPLC determination of tacrolimus and its related substances[J]. Chin J Pharm Anal, 2008, 28(2):309
[13] 莎日娜, 张冬婕, 赵志刚, 等. UHPLC-MS/MS测定人全血他克莫司浓度的不确定度评定[J]. 中国药师, 2021, 24(9):1752
SHA RN, ZHANG DJ, ZHAO ZG, et al. Uncertainty evaluation for tacrolimus analysis in human whole blood by UHPLC-MS/MS[J]. China Pharm, 2021, 24(9):1752
[14] 夏路风, 薛峰, 刘蕾, 等. HPLC-MS/MS测定肝移植早期患者全血中他克莫司的浓度[J]. 药物分析杂志, 2010, 30(9):1672
XIA LF, XUE F, LIU L, et al. HPLC-MS/MS determination of blood tacrolimus concentrations in early liver transplant recipients[J]. Chin J Pharm Anal, 2010, 30(9):1672
[15] 秦伟, 王晓雪, 张相林, 等. 他克莫司浓度在移植患者全血、血浆与血细胞中的相关性研究[J]. 中国药房, 2019, 30(15):2105
QIN W, WANG XX, ZHANG XL, et al. Correlation of tacrolimus concentrations among transplant patients’ whole blood, plasma and blood cells[J]. China Pharm, 2019, 30(15):2105
[16] MEI S, WANG J, CHEN D, et al. Simultaneous determination of cyclosporine and tacrolimus in human whole blood by ultra-high performance liquid chromatography tandem mass spectrometry and comparison with a chemiluminescence microparticle immunoassay[J]. J Chromatogr B Analyt Technol Biomed Life Sci, 2018, 1087-1088: 36
