Micelle-mediated extraction of tanshinones from Salvia miltiorrhiza bunge with analysis by high-performance liquid chromatography

The feasibility of employing non-ionic surfactant oligoethylene glycol monoalkyl ether (Genapol X-080) as an alternative and effective solvent for the extraction of tanshinones from Salvia miltiorrhiza bunge was studied for the first time. Various experimental conditions were investigated to optimize the extraction. Under optimum conditions, i.e. 10% Genapol X-080 (w/v), liquid/solid ratio of 20:1 (ml g⁻¹), ultrasonic-assisted extraction for 45 min, the extraction recovery of the tanshinones reached the highest value. When compared with commonly used solvents, 10% Genapol X-080 yielded almost the same extraction efficiency as methanol and dichloromethane-methanol (1:4). For the pre-concentration of tanshinones by cloud-point extraction (CPE), sodium chloride was added to the solution to facilitate the phase separation and increase the pre-concentration factor by reducing the volume of the surfactant-rich phase.     

Quality evaluation of Salvia miltiorrhiza Bge. by ultra high performance liquid chromatography with photodiode array detection and chemical fingerprinting coupled with chemometric analysis

An ultra high performance liquid chromatography with photodiode array detection method is developed for the simultaneous quantitative determination of five water‐soluble compounds including danshensu, protocatechualdehyde, rosmarinic acid, salvianolic acid B, and salvianolic acid A in Salvia miltiorrhiza Bge. samples. Through method optimization, the five compounds all expressed good linearity (R² > 0.9990) in a wide concentration range together with satisfactory accuracy, precision, and stability. Moreover, through qualitative analysis of the chemical fingerprint combined with similarity analysis, hierarchical cluster analysis, principle component analysis, and partial least‐squares discriminate analysis, we determined that the 13 batches of Salvia miltiorrhiza Bge. were similar in internal quality and the differences resulted from various cultivation environments, recovery elements, and others. Seen from the results of hierarchical cluster analysis and principle component analysis, the classification of 13 batches was in accordance, and partial least‐squares discriminate analysis technique was more suitable than the principle component analysis model to provide a distinct classification of test samples on the basis of their different components. Moreover, a permutation test verified the rationality of partial least‐squares discriminate analysis and variable importance plot showed that peaks 37 and 38 were the most significant variables in distinguishing the Salvia miltiorrhiza Bge. samples. The idea of the quantitative and qualitative analysis of Salvia miltiorrhiza Bge. was convenient, sensitive, and comprehensive, which could be applied to evaluate the quality of more traditional Chinese medicines.     

Fingerprint analysis,multi‐component quantitation,and antioxidant activity for the quality evaluation of Salvia miltiorrhiza var. alba by high‐performance liquid chromatography and chemometrics

Salvia miltiorrhiza Bge. var. alba C.Y. Wu and H.W. Li has wide prospects in clinical practice. A useful comprehensive method was developed for the quality evaluation of S. miltiorrhiza var. alba by three quantitative parameters: high‐performance liquid chromatography fingerprint, ten‐component contents, and antioxidant activity. The established method was validated for linearity, precision, repeatability, stability, and recovery. Principal components analysis and hierarchical clustering analysis were both used to evaluate the quality of the samples from different origins. The results showed that there were category discrepancies in quality of S. miltiorrhiza var. alba samples according to the three quantitative parameters. Multivariate linear regression was adopted to explore the relationship between components and antioxidant activity. Three constituents, namely, danshensu, rosmarinic acid, and salvianolic acid B, significantly correlated with antioxidant activity, and were successfully elucidated by the optimized multivariate linear regression model. The combined use of high‐performance liquid chromatography fingerprint analysis, simultaneous multicomponent quantitative analysis, and antioxidant activity for the quality evaluation of S. miltiorrhiza var. alba is a reliable, comprehensive, and promising approach, which might provide a valuable reference for other herbal products in general to improve their quality control.     

Binary chromatographic fingerprinting for quality evaluation of Radix Ophiopogonis by high-performance liquid chromatography coupled with ultraviolet and evaporative light-scattering detectors

Radix Ophiopogonis is a widely used traditional Chinese medicine. The quality of Radix Ophiopogonis available in the market varies, and some confusing or fake herbs exist. In order to improve the quality control of Radix Ophiopogonis, a novel fingerprinting method was established using HPLC coupled with UV and evaporative light-scattering detectors (ELSDs). Extraction with methanol and liquid-liquid extraction with water-saturated n-butanol were employed for the preparation of the sample solution. Chromatographic separation was performed on a Lichrospher C(18) column (250x4.6 mm id, 5.0 microm particle size) with a linear gradient elution program. UV detection at 280 nm and evaporative light-scattering detection were utilized to obtain two subfingerprinting chromatograms. A novel protocol for data processing was proposed, in order to identify and remove redundant data obtained by the two detectors, and balance the weight of the two subfingerprints on the similarity values. The method was validated and applied to quality evaluation of 16 samples of Radix Ophiopogonis and related herbs.     

Principal component analysis versus fuzzy principal component analysis A case study: the quality of danube water (1985-1996)

Principal component analysis (PCA) is a favorite tool in environmetrics for data compression and information extraction. PCA finds linear combinations of the original measurement variables that describe the significant variations in the data. However, it is well-known that PCA, as with any other multivariate statistical method, is sensitive to outliers, missing data, and poor linear correlation between variables due to poorly distributed variables. As a result data transformations have a large impact upon PCA. In this regard one of the most powerful approach to improve PCA appears to be the fuzzification of the matrix data, thus diminishing the influence of the outliers. In this paper we discuss and apply a robust fuzzy PCA algorithm (FPCA). The efficiency of the new algorithm is illustrated on a data set concerning the water quality of the Danube River for a period of 11 consecutive years. Considering, for example, a two component model, FPCA accounts for 91.7% of the total variance and PCA accounts only for 39.8%. Much more, PCA showed only a partial separation of the variables and no separation of scores (samples) onto the plane described by the first two principal components, whereas a much sharper differentiation of the variables and scores is observed when FPCA is applied.     

微柱高效液相色谱法测定丹参中的几种有效成分

研究了用微柱高效液相色谱法测定丹参中的4种有效成分(丹参素,原儿茶酸,原儿茶醛,丹酚酸B)的方法。丹参样品中的几种有效成分用体积分数40%的甲醇超声振荡提取,然后以WatersXterraTMRP18(1.0×50mm,2.5μm)微柱为固定相,甲醇和体积分数1%的HAc溶液梯度洗脱为流动相分离,在该色谱条件下,丹参中的4种有效成分在4.0min内可达到基线分离;用紫外二极管矩阵检测器检测。方法标准回收率为97%~102%,相对标准偏差为1.6%~2.3%。用此方法可测定几种丹参样品中的有效成分。     

Simultaneous qualitation and quantification of four phytoecdysones in Radix Achyranthis Bidentatae by high-performance liquid chromatography with diode array detection

A high-performance liquid chromatographic method with diode array detection was developed and validated to simultaneously identify and quantify four phytoecdysones,i.e., polypodine B (1), ecdysterone (2), 25-R inokosterone (3) and 25-S inokosterone (4), in Radix Achyranthis Bidentatae. The analysis was performed using a C(18) column with 6% tetrahydrofuran aqueous and acetonitrile isocratic elution, and the detection was carried out by DAD at 242 nm. The identities of the analytes were determined by comparing the retention time and UV spectrum with those of reference compounds. The validation of the method included linearity, sensitivity, recovery and stability. All calibration curves of the four phytoecdysones showed good linear regression (r(2) >or= 0.9993). The limit of detection (S/N = 3) and limit of quantification (S/N = 10) were less than 7.5 and 12.3 ng, respectively. The method provided desirable repeatability with overall intra- and inter-day variations of less than 4.67%. The obtained recoveries varied between 95.1 and 104.4% while the relative standard deviations were below 4.85% (n = 3). The analysis results indicated that the contents of the investigated phytoecdysones in Radix Achyranthis Bidentatae from different locations were highly variant, and the genuine crude drug indigenous to Henan province did not possess the highest concentration of phytoecdysones.     

Determination of the lipophilicity of Salvia miltiorrhiza Radix et Rhizoma (danshen root) ingredients by microemulsion liquid chromatography: optimization using cluster analysis and a linear solvation energy relationship‐based method

We evaluated 26 microemulsion liquid chromatography (MELC) systems for their potential as high‐throughput screening platforms capable of modeling the partitioning behaviors of drug compounds in an n‐octanol–water system, and for predicting the lipophilicity of those compounds (i.e. logP values). The MELC systems were compared by cluster analysis and a linear solvation energy relationship (LSER)‐based method, and the optimal system was identified by comparing their Euclidean distances with the LSER coefficients. The most effective MELC system had a mobile phase consisting of 6.0% (w/w) Brij35 (a detergent), 6.6% (w/w) butanol, 0.8% (w/w) cyclohexane, 86.6% (w/w) buffer solution and 8 mm cetyltrimethyl ammonium bromide. The reliability of the established platform was confirmed by the agreement between the experimental data and the predicted values. The logP values of the ingredients of danshen root (Salvia miltiorrhiza Radix et Rhizoma) were then predicted. Copyright © 2015 John Wiley & Sons, Ltd.     

Simultaneous analysis of seven astragalosides in Radix Astragali and related preparations by liquid chromatography coupled with electrospray ionization time-of-flight mass spectrometry

A method has been developed for the qualitative and quantitative analysis of pharmacologically active astragalosides isolated from several species of the genus Astragalus by high performance liquid chromatography coupled with electrospray ionization time-of-flight mass spectrometry. Seven astragalosides in Radix Astragali and their commercial pharmaceutical preparations were analyzed using the developed method. The extracted ion current chromatograms were obtained from the total ion current chromatogram using the m/z of [M+Na]+ ions produced by target compounds for peak determination. The limits of detection and limits of quantification were in the range of 0.10-0.22 ng and 0.22-0.52 ng in full scan mode, respectively. All calibration curves showed good linear regression (r2 > or = 0.9965) within the test range. The overall intra- and inter-day precision was less than 2.86% for peak area and the accuracy was higher than 92.9% on using ginsenoside I as internal standard. The assay was successfully utilized to analyze the major biologically active astragalosides in six samples of Astragalus membranaceus (Fisch.) Bge var. mongholicus (Bge.) Hsiao. and eight commercial preparations. The overall results demonstrate that this method is simple, selective, and suitable for the quality control of Chinese medicine and their preparation in the low nanogram range.     

Preparative separation of lithospermic acid B from Salvia miltiorrhiza by polyamide resin and preparative high-performance liquid chromatography

Adsorption on polyamide resin was investigated as a means of separating lithospermic acid B (LAB) from a crude extract of the roots of the traditional Chinese medicine Salvia miltiorrhiza Bunge ("Danshen"). Variables affecting adsorption capacity (solution pH, contact time on resin, initial LAB concentration) were studied. Adsorption was strongly dependent upon the initial concentration of LAB and pH. In all conditions, the polyamide resin gave optimal adsorption of LAB at an initial concentration of 2.66 mg/mL and pH <3.0. The adsorption isotherm correlated well with the Langmuir-type adsorption isotherm. Maximal adsorption capacity was calculated to be 380 mg/g at pH 2.0 and 25°C. LAB purity of 85.30% could be obtained by polyamide resin adsorption followed by elution with 70% ethanol solution, and the recovery was 87.1%. After preparative HPLC, the maximum HPLC purity obtained was 99.28% with a recovery of 75.2%. This method provides an efficient and low-cost method for LAB purification for industrial applications.     

Quality evaluation of Radix Stemonae through simultaneous quantification of bioactive alkaloids by high-performance liquid chromatography coupled with diode array and evaporative light scattering detectors

A high-performance liquid chromatography coupled with diode array detection and evaporative light scattering detection (HPLC-DAD-ELSD) method was developed to simultaneously quantify six major bioactive alkaloids belonging to different structure types in Radix Stemonae, Bai-Bu in Chinese, a traditionally used antitussive and insecticidal medicinal material in China and other countries of Southeast Asia. Diode array detector (DAD) with the wavelengths at 307 and 260 nm was used to monitor the conjugated system of protostemonine (2) and maistemonine (4), respectively, whereas evaporative light scattering detector (ELSD) was employed to detect croomine (1), stemoninine (3), neotuberostemonine (5) and tuberostemonine (6), the other four analytes with no or poor chromophores. The assay was validated to be sensitive, precise and accurate, with a detection limit of 3.64-0.04 microg/mL depending on the individual analytes. The overall intra- and inter-day variations were less than 9.3%, and the overall recoveries higher than 91.2%, respectively. The correlation coefficients of the calibration curves were better than 0.996 for all analytes. The newly established method was successfully utilized to determine six major components in the genuine sources of Radix Stemonae: Stemona japonica, S. sessilifolia and S. tuberosa. Significant variations of contents of these components were demonstrated in samples of these three species. This simple, rapid, low-cost and reliable method is suitable for the routine quality control of herbal medicines containing bioactive components with different structure types such as Radix Stemonae.     

Hollow Fiber/Solvent Bar Microextraction Coupled with High Performance Liquid Chromatography for Preconcentration and Determination of Tanshinones and Salvianolic Acids in Radix Salvia miltiorrhiza

Hollow fiber solvent bar microextraction coupled with high-performance liquid chromatography was developed for the preconcentration and determination of active ingredients in Radix Salvia miltiorrhiza. These ingredients include dihydrotanshinone I, cryptotanshinone, tanshinone I, tanshinone IIA, salvianolic acid B, danshensu, and protocatechuic aldehyde. To evaluate the technique, seven compounds of varying polarity were used as model analytes, and a polyvinylidene fluoride hollow fiber (1.0 cm) with octanol (2 µL) was used as microextraction bar. The extraction conditions, including the identity of the hollow fiber, organic solvent, pH, salt addition, agitation speed, extraction time, and volume, were investigated and optimized. The extraction mechanism was analyzed and verified. The two main parameters, extraction recovery and enrichment factor, were obtained. Under the most favorable conditions, the enrichment factors of the analytes were 0.7–612, the limits of detection were below 1.11 ng mL^?1, and the recoveries were between 95.4% and 101.3%. Thus, hollow fiber solvent bar microextraction is simple, rapid, and practical with a wide range of potential applications.     

三波长融合指纹图谱结合6组分定量和主成分分析评价银翘解毒片的质量

建立了三波长融合高效液相色谱指纹图谱,并结合6组分定量和主成分分析(PCA)评价25批银翘解毒片的质量一致性。采用反相高效液相色谱法(RP-HPLC)分别于230、279、327 nm下检测。运用多波长融合指纹图谱技术建立银翘解毒片三波长融合指纹图谱,采用系统指纹定量法对其进行定性和整体定量评价。结果鉴别出25批银翘解毒片样品完全合格且区分良好。同时测定6组分含量,与指纹图谱结合,从整体和局部角度评价银翘解毒片质量。此外,运用PCA法对融合指纹图谱进行分析,通过主成分得分图可以明显区分来自两个厂家的25批银翘解毒片样品。方法综合性较强且有效,为科学评价与有效控制银翘解毒片的质量提供了可靠的参考。     

Thermogravimetric and principal component analyses in quality assessment of lubricating oils

The suitability of the derivative thermogravimetric and principal component analyses for the assessment of service performance of lubricating oils has been studied. A total sum of 179 samples has been examined, including M-20 Bp, MS-20 p, Marinol CB SAE-30 and DS-11 oils. The results indicate that principal component analysis greatly assisted in the analysis of the quality of lubricating oils by derivative thermogravimetric technique. Considering that, this multivariate statistical method can be applied to the differentiation of oil samples taking into account degree of their degradation in the oil system of an engine.     

基于主成分分析的骨碎补药材乙醇和环己烷提取物的高效液相色谱指纹图谱及指标成分的定量分析

建立了骨碎补药材乙醇和环己烷提取物的高效液相色谱(HPLC)指纹图谱,并利用主成分分析法(PCA)对指纹图谱进行统计分析,以各主要色谱峰的保留时间和峰面积为变量得到score图和loading图。在score图和loading图中,骨碎补的正品和非正品可明显区分,且揭示出对此区分贡献最大的4个潜在指标成分,其中已知成分为柚皮苷、新北美圣草苷和E-4-O-β-D-葡萄糖酰咖啡酸。同时测定了这3种成分在19批正品和非正品骨碎补药材中的含量,其中10批骨碎补药材正品中3种成分的含量为: 柚皮苷6.36～10.1 mg/g,新北美圣草苷5.14～9.21 mg/g,E-4-O-β-D-葡萄糖酰咖啡酸1.87～3.19 mg/g。该方法更全面地反映了药材的化学成分信息,并能从定性和定量两方面控制骨碎补药材的内在质量。     

Simultaneous quantification of seven bioactive components in Caulis Lonicerae Japonicae by high performance liquid chromatography

This study presents a new HPLC method for the simultaneous determination of seven major components, namely chlorogenic acid, caffeic acid, loganin, sweroside, secoxyloganin, rutin and luteolin 7-O-glucoside in Caulis Lonicerae Japonicae, a commonly used traditional Chinese medicinal herb derived from the caulis of Lonicera japonica Thunb. These seven compounds, belonging to the chemical types of phenolic acids, iridoids and flavonoids, were separated on a C18 column (250 x 4.6 mm, 5.0 microm) with the column temperature at 30 degrees C. The mobile phase was composed of (A) aqueous acetic acid (0.4%, v/v) and (B) acetonitrile using a gradient elution of 10% B at 0-12 min, 10-17% B at 12-25 min and 17% B at 25-35 min. The flow rate was 1.0 mL/min and detection wavelength was set at 245 nm. The limit of detection (S/N = 3) ranged from 0.10 to 0.23 microg/mL and the limit of quantification (S/N = 10) ranged from 0.69 to 3.56 microg/mL. All calibration curves showed good linear regression (r2 > 0.9990) within the test ranges. The intra- and inter-day precisions as determined from sample solutions were below 1.24 and 2.28%, respectively. The recoveries for seven compounds were found to range from 94.2 to 103.6%. This verified method has been successfully applied to evaluation of commercial samples of Caulis Lonicerae Japonicae from different markets in China.     

Determination of fifteen bioactive components in Radix et Rhizoma Salviae Miltiorrhizae by high-performance liquid chromatography with ultraviolet and mass spectrometric detection

A high-performance liquid chromatographic (HPLC) method coupled with ultraviolet (UV) and electrospray ionization time-of-flight mass spectrometry (ESI-TOF/MS) was established for simultaneous qualitative and quantitative determination of nine phenolic acids and six diterpenoids in Radix et Rhizoma Salviae Miltiorrhizae (RRSM). The optimal chromatographic conditions were achieved on a Zorbax C(18) column by gradient elution with 0.1% (v/v) aqueous formic acid and acetonitrile as mobile phase at the flow rate of 1.0 mL/min. The detection wavelength at 281 nm was chosen to determine the 15 bioactive components, namely danshensu (1), protocatechuic acid (2), protocatechuic aldehyde (3), caffeic acid (4), rosmarinic acid (5), lithospermic acid (6), salvianolic acid B (7), salvianolic acid A (8), salvianolic acid C (9); dihydrotanshinone I (10), cryptotanshinone (11), tanshinone I (12), methylene tanshiqunone (13), tanshinone IIA (14) and miltirone (15). Additionally, LC-ESI-TOF/MS was used to make definite identification of the constituents in samples in comparison with those reference compounds. The validation of the method included tests of linearity, sensitivity, repeatability, stability and recovery. The proposed method was successfully applied to quantify the 15 components in 21 samples; significant variations were demonstrated in the contents of the samples from diverse species and origins. The developed method could be used to effectively and comprehensively evaluate the quality of RRSM for its clinical safety and efficacy.     

Investigation of sport supplements quality by Raman spectroscopy and principal component analysis

In this work, sport supplements were investigated by Raman spectroscopy. Samples were obtained from health foods shops, gyms and sports centers covering a wide range of available supplement powders. A systematic comparison of Raman spectra of the analyzed supplements allowed identifying the supplement type through the characteristic vibrational modes of carbohydrates and proteins. The protein supplements were identified by Raman bands at 1650, 1250 and 1004 cm⁻¹, while the spectral range between 1200 and 800 cm⁻¹ was useful to identify the carbohydrate supplements. Due to the diversity in composition of sport supplements, a chemometric tool such as principal component analysis (PCA) was employed to assist in the interpretation of Raman spectra, allowing also the identification of compounds present in sport supplements. Especially, the Raman scattering of aromatic and aliphatic amino acids residues contributes to the existence of bands characteristic for the different types of proteins. This kind of information is very important for the quality control of these products, for detecting the presence of fraud or a sample composition in disagreement with the label, thus ensuring the provenance of the supplements.