高效液相色谱法测定化妆品中7种甘草成分

建立了一种测定化妆品中7种甘草成分的高效液相色谱法。样品经甲醇(7+3)溶液提取,采用ODS-2HYPERSIL色谱柱进行分离,以乙腈-0.05%(φ)磷酸溶液为流动相进行梯度洗脱,采用光电二极管阵列检测器在237nm处进行检测。7种组分的质量浓度在1~200mg·L-1范围内与其峰面积呈线性关系,检出限(3S/N)在0.6~1.0ng之间。对空白样品进行加标回收试验,回收率在92.5%~103%之间,测定值的相对标准偏差(n=6)在0.09%~1.8%之间。     

Isolation and purification of inflacoumarin A and licochalcone A from licorice by high-speed counter-current chromatography

High-speed counter-current chromatography (HSCCC) technique in semi-preparative scale has been applied to isolate and purify bioactive flavone compounds from the ethanol extract of Glycyrrhiza inflata Bat., a particular plant species of licorice. HSCCC separation was performed with a two-phase solvent system composed of n-hexane-chloroform-methanol-water (5:6:3:2, v/v) by eluting the lower mobile phase at a flow rate of 1.8 ml/min and a revolution speed of 800 rpm. Purification was performed with a two-phase solvent system composed of n-hexane-chloroform-methanol-water (1.5:6:3:2, v/v) by eluting the lower mobile phase at a flow-rate of 1.5 ml/min and a revolution speed of 800 rpm. Two major flavone peaks: inflacoumarin A and licochalcone A were collected and the respective yields of the peaks amount to 6 mg (8.6%, w/w) and 8 mg (11.4%, w/w) from 70 mg of the crude extract sample. The purities of inflacoumarin A and licochalcone A reached 99.6% and 99.1%, respectively, after a sequential purification run. The structures of inflacoumarin A and licochalcone A were positively confirmed by 1H NMR and 13C NMR, 1H-13C-COSY, UV, FT-IR and electron ionization MS analyses.     

Determination of ochratoxin A in licorice root using inverse ion mobility spectrometry

Despite the recent, successful efforts to detect mycotoxins, new methods are still required to achieve higher sensitivity, more simplicity, higher speed, and higher accuracy at lower costs. This paper describes the determination of ochratoxin A (OTA) using corona discharge ion mobility spectrometry (IMS) in the licorice root. A quick screening and measuring method is proposed to be employed after cleaning up the extracted OTA by immunoaffinity columns. The ion mobility spectrometer is used in the inverse mode to better differentiate the OTA peak from the neighboring ones. After optimization of the experimental conditions such as corona voltage, injection port temperature, and IMS cell temperature, a limit of detection (LOD) of 0.010 ng is obtained. Furthermore, the calibration curve is found to be in the range of 0.01-1 ng with a correlation coefficient (R²) of 0.988. Licorice roots were analyzed for their OTA content to demonstrate the capability of the proposed method in the quantitative detection of OTA in real samples.     

甘草中金属元素与总黄酮含量测定与抗氧化研究

对中草药甘草中金属元素与总黄酮含量进行测定,并对其清除超氧自由基进行研究.测定结果显示,甘草中含有丰富的金属元素和黄酮类化合物,其中总黄酮为16.42%,金属元素含量为铁>锌>锰>铜,对超氧自由基具有一定的清除作用.实验结果为研究甘草治疗呼吸系统疾病及清除自由基提供了有用的数据.     

复杂中药体系色谱分离过程中进样条件的优化与预测

以甘草提取物为原料,甘草苷为目标组分,对复杂中药体系色谱分离过程中的进样条件进行优化。以70%产品纯度下甘草苷的回收率y为目标函数,以进样纯度x1和进样浓度x2为自变量,提出4种模型,并采用计算模拟及回归分析的方法得出模型参数,得到复杂甘草提取物色谱进样条件的非线性模型为y=63.61x10.204x2-0.09。结合分析预测结果,对甘草苷进行了分离纯化。     

Identification and quantification of anticarcinogens in garlic extract and licorice root extract powder

Nine organosulfur compounds present in an aged garlic extract and two isoflavonoids and one triterpenoid present in a licorice root extract powder have been identified and quantified. Quantification involved solvent extraction and gas chromatographic – mass spectrometric analysis (garlic extract) or hydrolysis, solvent extraction, and liquid chromatographic analysis (licorice root extract powder). Although the garlic extract proved to be unstable and the concentration of the organosulfur compounds varied with time, one analysis of the extract gave the following results: methyl disulfide (0.607 μg/g), methyl trisulfide (0.181 μg/g), allyl sulfide (2.02 μg/g), allyl disulfide (0.784 μg/g), allyl trisulfide (0.795 μg/g), allyl methyl sulfide (1.64 μg/g), allyl methyl disulfide (0.411 μg/g), allyl methyl trisulfide (0.695 μg/g), and ethyl 2-propenesulfinate (11.4 μg/g). The analysis of the licorice root extract powder gave the following results: formononetin (1.92 mg/g), isoliquiritigenin (9.61 mg/g), and 18β-glycyrrhetinic acid (43.9 mg/g). Methods were successfully developed to quantify the same compounds in the serum of test animals which had consumed feed spiked with garlic extract or licorice root extract powder. Only the 18β-glycyrrhetinic acid could be detected in the sera of such animals, however. An effort was also made to determine serum levels of prostaglandin E2 to correlate its inhibition with levels of the dietary components, but the prostaglandin E2 levels were too low to measure.     

Combination of the advantages of chromatographic methods based on active components for the quality evaluation of licorice

A rapid, improved and comprehensive method including high‐performance thin‐layer chromatography, fingerprint technology and single standard to determine multiple components was developed and validated for the quality evaluation of licorice. In this study, a newly developed high‐performance thin‐layer chromatography method was first used for authentication of licorice, which achieved simultaneous identification of multiple bands including five bands for known bioactive components by comparing their retention factor values and colors with the standards. For fingerprint analysis, 8 of 16 common peaks were identified. Simultaneously, similarity analysis which showed very similar patterns and hierarchical clustering analysis were performed to discriminate and classify the 27 batches of samples. Additionally, the single standard to determine multiple components method was first successfully achieved to quantify the eight important active markers in licorice including liquiritin apioside, liquiritin, isoliquiritin apioside, isoliquritin, neoisoliquiritin, liquiritigenin, isoliquiritigenin and glycyrrhizic acid. The easily available glycyrrhizic acid was selected as the reference substance to calculate relative response factors. Compared with the normal external standard method, this alternative method can be used to determine the multiple indices effectively and accurately. The validation result showed that the developed method was specific, accurate, precise, robust and reliable for the overall quality assessment of licorice.     

Metabolite profiling of licorice (Glycyrrhiza glabra) from different locations using comprehensive two-dimensional liquid chromatography coupled to diode array and tandem mass spectrometry detection

Profiling of the main metabolites from several licorice (Glycyrrhiza glabra) samples collected at different locations is carried out in this work by using comprehensive two-dimensional liquid chromatography (LC × LC) coupled to diode array (DAD) and mass spectrometry (MS) detectors. The optimized method was based on the application of a HILIC-based separation in the first dimension combined with fast RP-based second dimension separation. This set-up was shown to possess powerful separation capabilities allowing separating as much as 89 different metabolites in a single sample. Identification and grouping of metabolites according to their chemical class were achieved using the DAD, MS and MS/MS data. Triterpene saponins were the most abundant metabolites followed by glycosylated flavanones and chalcones, whereas glycyrrhizic acid, as expected, was confirmed as the main component in all the studied samples. LC × LC-DAD-MS/MS was able to resolve these complex licorice samples providing with specific metabolite profiles to the different licorice samples depending on their geographical origin. Namely, from 19 to 50 specific compounds were exclusively determined in the 2D-chromatograms from the different licorice samples depending on their geographical origin, which can be used as a typical pattern that could potentially be related to their geographical location and authentication.     

Purification of flavonoids from licorice using an off‐line preparative two‐dimensional normal‐phase liquid chromatography/reversed‐phase liquid chromatography method

An orthogonal (71.9%) off‐line preparative two‐dimensional normal‐phase liquid chromatography/reversed‐phase liquid chromatography method coupled with effective sample pretreatment was developed for separation and purification of flavonoids from licorice. Most of the nonflavonoids were firstly removed using a self‐made Click TE‐Cys (60 μm) solid‐phase extraction. In the first dimension, an industrial grade preparative chromatography was employed to purify the crude flavonoids. Click TE‐Cys (10 μm) was selected as the stationary phase that provided an excellent separation with high reproducibility. Ethyl acetate/ethanol was selected as the mobile phase owing to their excellent solubility for flavonoids. Flavonoids co‐eluted in the first dimension were selected for further purification using reversed‐phase liquid chromatography. Multiple compounds could be isolated from one normal‐phase fraction and some compounds with bad resolution in one‐dimensional liquid chromatography could be prepared in this two‐dimensional system owing to the orthogonal separation. Moreover, this two‐dimensional liquid chromatography method was beneficial for the preparation of relatively trace flavonoid compounds, which were enriched in the first dimension and further purified in the second dimension. Totally, 24 flavonoid compounds with high purity were obtained. The results demonstrated that the off‐line two‐dimensional liquid chromatography method was effective for the preparative separation and purification of flavonoids from licorice.     

Ultra high performance liquid chromatography coupled with triple quadrupole mass spectrometry and chemometric analysis of licorice based on the simultaneous determination of saponins and flavonoids

Licorice is among the most popular herbal medicines and frequently used in traditional medicine, food products, and cosmetics. In China, only Glycyrrhiza uralensis Fisch., Glycyrrhiza inflata Bat. and Glycyrrhiza glabra L. are officially used and are usually processed with honey prior to use. To maintain the quality of commercially available herbal products, a simple, rapid, and reliable ultra high performance liquid chromatography with triple quadrupole mass spectrometry was developed to investigate the major active constituents of commercially available licorice products. Nineteen components were accurately determined, including eight triterpenoid saponins, one triterpene, and ten flavonoids. Subsequently, multivariate statistical analysis methods were employed to further explore and interpret the experimental data. The results indicated that liquiritin apioside may be considered as a candidate index for the quality control of licorice as well as 18β‐glycyrrhizic acid and liquiritin. In addition, both 18β‐glycyrrhizic acid and licorice‐saponin G2 can be used for discrimination between crude and honey‐processed licorice. Furthermore, using 18β‐glycyrrhizic acid and liquiritin as markers, this work revealed that the quality of licorice products may have declined in recent years. This highlights the need for additional effort focused on good agricultural practice during the processing of licorice. In summary, this study provides a valuable reference for the quality assessment of licorice.