目的: 采用高分辨质谱及组学分析技术分析狭叶番泻叶及耳叶番泻叶化学成分差异,并建立狭叶番泻叶中掺伪耳叶番泻叶的快速检查方法。方法: 采用超高效液相色谱-四极杆飞行时间串联质谱(UPLC-Q TOF MS),分别采集13批狭叶番泻叶和9批耳叶番泻叶的正谱和负谱MSE数据。采用Agilent ZORBAX SB-C18(100 mm×2.1 mm,1.8 μm)色谱柱,柱温30 ℃,以乙腈(A)-1%乙酸(B)为流动相,流速0.3 mL·min-1,进样体积1 μL;采集质量范围为m/z 50~1 200。运用组学分析软件QI对狭叶番泻叶和耳叶番泻叶负谱数据进行正交偏最小二乘法判别分析,筛选出7个耳叶番泻叶特征性成分,并对其中的1个主要特征性成分进行了分离鉴定。进一步以主要特征性成分为指标性成分建立了快速检查狭叶番泻叶中掺伪耳叶番泻叶的UPLC方法,并应用于27批狭叶番泻叶样品及3批自制阳性样品的测定。结果: 狭叶番泻叶和耳叶番泻叶的化学成分差异显著,经鉴定耳叶番泻叶中特有成分为kaempferol 3-O-(2”-O-apiofuranosyl) rutinoside。在狭叶番泻叶中掺伪耳叶番泻叶的UPLC检查方法中,该特有成分的精密度(RSD=1.3%)、重复性(RSD=1.3%)和稳定性(RSD=0.58%)均符合要求。27批狭叶番泻叶样品中23批未检出kaempferol 3-O-(2”-O-apiofuranosyl)rutinoside;4批狭叶番泻叶样品及3批自制阳性样品中检出kaempferol 3-O-(2”-O-apiofuranosyl)rutinoside。结论: 本研究应用UPLC-Q TOF MS技术结合OPLS-DA分析方法成功将狭叶番泻叶和耳叶番泻叶进行了区分,分离并鉴定出耳叶番泻叶的主要特征性成分,并建立了快速筛查狭叶番泻叶中掺伪耳叶番泻叶的UPLC方法,为狭叶番泻叶的质量控制及质量标准的完善提供了依据,为中药材掺伪鉴别研究提供了思路和方法。
Objective: To evaluate the chemical composition difference of Cassia angustifolia Vahl leaves and Cassia auriculata L. leaves by high-resolution mass spectrometry and omics analysis, so as to establish the detection method of Cassia auriculata L. leaves adulterated in Cassia angustifolia Vahl leaves. Methods: The positive and negative MSE data of 13 batches of Cassia angustifolia Vahl leaves and 9 batches of Cassia auriculata L. leaves were collected by ultra performance liquid chromatography coupled with electrospray ionization quadrupole time of flight mass spectrometry(UPLC-Q TOF MS). The mobile phase was acetonitrile(A)-1% acetic acid(B). The column temperature was 30 ℃. The flow rate was 0.3 mL·min-1. Injection volume was 1 μL. The mass range was m/z 50-1 200. The chemical composition difference of Cassia angustifolia Vahl leaves and Cassia auriculata L. leaves were processed by the omics analysis QI software based on the orthogonal partial least-squares discrimination analysis(OPLS-DA) after the negative MSE data were obtained. One out of seven specific components from Cassia auriculata L. leaves was separated and identified. A method with the specific component as the reference substance for the detection of adulterated Cassia auriculata L. leaves in Cassia angustifolia Vahl leaves was established by UPLC, and it was used in 27 batches of Cassia angustifolia Vahl leaves samples and 3 batches of laboratory-made positive samples. Results: Cassia angustifolia Vahl leaves and Cassia auriculata L. leaves were significantly different from each other. The specific component separated from Cassia auriculata L. leaves was identified as kaempferol 3-O-(2”-O-apiofuranosyl) rutinoside. In the method for the detection of adulterated Cassia auriculata L. leaves in Cassia angustifolia Vahl leaves by UPLC, the precision(RSD=1.3%), repeatability(RSD=1.3%) and stability(RSD=0.58%) met the requirements. No kaempferol 3-O-(2”-O-apiofuranosyl) rutinoside was detected in 23 out of 27 batches of Cassia angustifolia Vahl leaves samples, but it was detected in 4 batches of Cassia angustifolia Vahl leaves samples and 3 batches of positive samples made in the laboratory. Conclusion: In this study, the difference of Cassia angustifolia Vahl leaves and Cassia auriculata L. leaves is distinguished clearly based on the technology of UPLC-Q TOF MS and OPLS-DA. The specific component of Cassia auriculata L. leaves is separated and identified. A method for the detection of adulterated Cassia auriculata L. leaves in Cassia angustifolia Vahl leaves is established by UPLC, and it provides the basis for quality control and quality standard improvement of Cassia angustifolia Vahl leaves. The technology is helpful to solve the problem of adulteration identification of traditional Chinese medicine.
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