超高效液相色谱-串联质谱法测定化妆品中新型糖皮质激素氯倍他索乙酸酯

doi:10.3724/SP.J.1123.2022.06010

摘要/Abstract

摘要：

采用超高效液相色谱-串联质谱法建立了化妆品中氯倍他索乙酸酯的检测方法,适用于膏霜乳类、凝胶类、泥类、贴膜类和液体(水)类5种常见化妆品基质。对影响被测物质的前处理方法、提取溶剂、提取时间等因素和色谱、质谱条件等进行了考察。最终建立方法为样品经乙腈涡旋分散、超声提取、过滤后用超高效液相色谱-串联质谱仪测定,采用Waters CORTECS C₁₈色谱柱(150 mm×2.1 mm, 2.7 μm),以水和乙腈作为流动相分离,在电喷雾正离子模式(ESI⁺)下,以多反应监测(MRM)方式采集,采用基质匹配标准曲线进行定量分析。5种基质类型下,被测物质在0.9~37 μg/L的范围内线性拟合良好,线性相关系数(R²)均超过0.99。方法的检出限为0.03 μg/g,定量限为0.09 μg/g。在定量限、2倍定量限和10倍定量限3个水平下的加标回收率试验中,被测物质在5种基质中的回收率范围为83.2%~103.2%,相对标准偏差(RSD, n=6)范围为1.4%~5.6%。采用该方法对不同基质类型的化妆品样品进行筛查,共发现5批阳性样品,其中氯倍他索乙酸酯的含量范围为1.1~48.1 μg/g。该方法操作简单,灵敏可靠,适用于不同基质类型化妆品的高通量定性定量筛查分析,为监测化妆品中的非法添加提供了技术支持及理论依据。

关键词: 超高效液相色谱-串联质谱, 糖皮质激素, 氯倍他索乙酸酯, 化妆品

Abstract:

At present, new prohibited substances are becoming more common illegal additions in cosmetics. Clobetasol acetate is a new glucocorticoid, which is not covered in the current national standards and is a homologue of clobetasol propionate. A method was established for the determination of clobetasol acetate as a new glucocorticoid (GC) in cosmetics by ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Five common cosmetic matrices were suitable for this new method: creams, gels, clay masks, masks and lotions. Four pretreatment methods were compared: direct extraction by acetonitrile, PRiME pass-through column purification, solid-phase extraction (SPE) purification, and QuEChERS purification. Further, the effects of different extraction efficiencies of the target compound, such as extraction solvents and extraction time, were investigated. The MS parameters, such as ion mode, cone voltage and collision energy of ion pairs of the target compound, were optimized. The chromatographic separation conditions and response intensities of the target compound in different mobile phases were compared. Based on the experimental results, the optimal extraction method was determined to be direct extraction, wherein the samples were vortex dispersed with acetonitrile, ultrasonic extraction over 30 min and filtered by a 0.22 μm organic millipore filter, and then the samples were detected by UPLC-MS/MS. The concentrated extracts were separated on a Waters CORTECS C₁₈ column (150 mm×2.1 mm, 2.7 μm), where the water and acetonitrile were used as the mobile phases for gradient elution. The target compound was detected with the multiple reaction monitoring (MRM) mode under electrospray ionization and positive ion scanning (ESI⁺). Quantitative analysis was performed by matrix matching standard curve. Under the optimum conditions, the target compound had good linear fitting in the range of 0.9-37 μg/L. The linear correlation coefficient (R²) was greater than 0.99, the limit of quantification (LOQ) of the method was 0.09 μg/g and the limit of detection (LOD) was 0.03 μg/g for these five different cosmetic matrices. The recovery test was conducted under three spiked levels: 1, 2 and 10 times of LOQ. The recoveries of the tested substance were between 83.2% and 103.2% in these five cosmetic matrices, and the relative standard deviations (RSDs, n=6) were between 1.4% and 5.6%. This method was used to screen cosmetic samples of different matrix types, and a total of five positive samples were found, in which the content range of clobetasol acetate was from 1.1 to 48.1 μg/g. In conclusion, the method is simple, sensitive and reliable, and is suitable for high-throughput qualitative and quantitative screening, and the analysis of cosmetics with different matrix types. Moreover, the method provides crucial technical support and a theoretical basis for the establishment of feasible detection standards for clobetasol acetate in China, as well as for the control of the compound in cosmetics. This method has important practical significance to implement management measures of illegal additions in cosmetics.

Key words: ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS), glucocorticoids, clobetasol acetate, cosmetics

中图分类号:

O658

杨飘飘, 黄伟, 李丽霞, 刘红. 超高效液相色谱-串联质谱法测定化妆品中新型糖皮质激素氯倍他索乙酸酯[J]. 色谱, 2023, 41(3): 250-256.

YANG Piaopiao, HUANG Wei, LI Lixia, LIU Hong. Determination of new glucocorticoid called clobetasol acetate in cosmetics by ultra performance liquid chromatography-tandem mass spectrometry[J]. Chinese Journal of Chromatography, 2023, 41(3): 250-256.

图/表 9

图1 氯倍他索乙酸酯的结构

Fig. 1 Structural formula of clobetasol acetate

表1 氯倍他索乙酸酯的保留时间和质谱参数

Table 1 Retention time and MS parameters for clobetasol acetate

Compound	Retention time/ min	Parent ion (m/z)	Quantitative/ qualitative ions (m/z)	Cone voltage/ V	Collison energies/ V
Clobetasol	4.67	453.2	278.2/263.2	24	24/28
acetate

图2 目标化合物在不同样品前处理方法下的提取回收率(n=6)

Fig. 2 Extraction recoveries of the target compound under different sample pretreatment methods (n=6) ①: extraction by acetonitrile; ②: solid-phase extraction; ③: PRiME pass-through column purification; ④: QuEChERS purification.

图3 目标化合物在不同提取溶剂条件下的提取回收率(n=6)

Fig. 3 Extraction recoveries of the target compound under different extraction solvents (n=6) ①: acetonitrile; ②: acetonitrile-water (70∶30, v/v); ③: acetonitrile-water (50∶50, v/v); ④: methanol; ⑤: methanol-water (70∶30, v/v); ⑥: methanol-water (50∶50, v/v).

图4 目标化合物在不同超声时间下的提取回收率(n=6)

Fig. 4 Extraction recoveries of the target compound under different ultrasonic times (n=6)

表2 在不同基质类型下目标化合物的线性范围、线性方程、相关系数、检出限、定量限及基质效应

Table 2 Linear ranges, linear equations, correlation coefficients (R2), limits of detection (LODs), limits of quantification (LOQs), and matrix effects (MEs) of the target compound under different matrix types

Matrix type	Linear range/(μg/L)	Linear equation	R²	LOD/(μg/g)	LOQ/(μg/g)	ME/%
Standard solution	0.9-37	y=2.57×10³x+6.39×10²	0.9998	0.03	0.09	/
Cream	0.9-37	y=1.34×10³x+5.04×10²	0.9982	0.03	0.09	52.2
Gel	0.9-37	y=8.73×10²x+2.17×10²	0.9958	0.03	0.09	34.0
Clay masks	0.9-37	y=1.89×10³x+9.13×10²	0.9984	0.03	0.09	73.7
Mask	0.9-37	y=1.95×10³x+9.02×10²	0.9998	0.03	0.09	76.0
Lotion	0.9-37	y=2.15×10³x+1.08×10³	0.9994	0.03	0.09	83.8

表3 不同基质类型下目标化合物的加标回收率和精密度(n=6)

Table 3 Spiked recoveries and precisions of the target compound in different matrix types (n=6)

Compound	Matrix type	0.09 μg/g		0.19 μg/g		0.94 μg/g
Compound	Matrix type	Recovery/%	RSD/%	Recovery/%	RSD/%	Recovery/%	RSD/%
Clobetasol	cream	94.4	3.3	96.2	2.2	101.1	5.6
acetate	gel	85.5	2.4	100.4	3.8	95.6	4.4
	clay mask	94.0	5.3	91.9	4.4	92.4	1.4
	mask	91.3	3.8	100.1	3.9	103.2	3.6
	lotion	83.2	3.9	94.8	3.2	96.5	1.4

表4 阳性样品的样品信息和含量

Table 4 Information and contents of positive samples

No.	Matrix type	Manufacturer	Sampling site	Content/ (μg/g)
1	mask	manufacturer 1	small shop	1.1
2	mask	manufacturer 1	small shop	1.1
3	mask	manufacturer 2	small shop	34.5
4	gel	manufacturer 3	online shopping	39.2
5	gel	manufacturer 3	production unit	48.1

图5 (a)典型阳性样品和(b)对照溶液中氯倍他索乙酸酯的色谱图

Fig. 5 Chromatograms of clobetasol acetate in (a) typical positive and (b)reference solution

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