State-of-the-art separation of ginsenosides from Korean white and red ginseng by countercurrent chromatography

Ginseng (Panax ginseng C. A. Meyer) has been one of the most popular herbs used for nutritional and medicinal purposes by the people of eastern Asia for thousands of years. Ginsenosides, the mostly widely studied chemical components of ginseng, are quite different depending on the processing method used. A number of studies demonstrate the countercurrent chromatography (CCC) separation of ginsenosides from several sources; however, there is no single report demonstrating a one-step separation of all of these ginsenosides from different sources. In the present study, we have successfully developed an efficient CCC separation methodology in which the flow-rate gradient technique was coupled with a new solvent gradient dilution strategy for the isolation of ginsenosides from Korean white (peeled off dried P. ginseng) and red ginseng (steam-treated P. ginseng). The crude samples were initially prepared by extraction with butanol and were further purified with CCC using solvent gradients composed of methylene chloride–methanol–isopropanol–water (different ratios, v/v). Gas chromatography coupled with flame ionization detector was used to analyze the components of the two-phase solvent mixture. Each phase solvent mixture was prepared without presaturation, which saves time and reduces the solvent consumption. Finally, 13 ginsenosides have been purified from red ginseng with the new technique, including Rg₁, Re, Rf, Rg_2, Rb₁, Rb₂, Rc, Rd, Rg₃, Rk₁, Rg₅, Rg₆, and F₄. Meanwhile, eight ginsenosides have been purified from white ginseng, including Rg₁, Re, Rf, Rh_1, Rb₁, Rb₂, Rc, and Rd by using a single-solvent system. Thus, the present technique could be used for the purification of ginsenosides from all types’ ginseng sources. To our knowledge, this is the first report involving the separation of ginsenoside Rg₂ and Rg₆ and the one-step separation of thirteen ginsenosides from red ginseng by CCC.     

Performance Evaluation of Charged Aerosol and Evaporative Light Scattering Detection for the Determination of Ginsenosides by LC

A sensitive LC-CAD method was developed for simultaneous determination of seven major triterpenoid saponins, namely ginsenosides Rg₁, Re, Rb₁, Rc, Rb₂, Rb₃ and Rd in Panax ginseng C. A. Meyer, a commonly used traditional Chinese medicine. This CAD method was evaluated in sensitivity, linearity and reproducibility compared to ELSD and UV. It was found the developed method has improved sensitivity, linearity and reproducibility compared to ELSD. This method was successfully applied to analyze the ginsenosides in ten samples of Panax ginseng. The validation results indicated that the improved method can be utilized as another approach for quality control of P. ginseng.     

Mechanism of Incompatible Herb Pairs,Panax ginseng and Veratrum nigrum L.: Material Basis and Metabolic Profiles of Ginsenosides in Rat Intestinal Bacteria

In traditional Chinese medicine theory, Panax ginseng and Veratrum nigrum L. is an important incompatible herb pair. Studies on the content variation of main components and the influences on the metabolism in rat intestinal bacteria are useful to understand the mechanism of incompatibility of this herb pairs. In this study, the content variation of ginsenosides and their metaboltic profiles in the extracts of P. ginseng and compatibility of P. ginseng with V. nigrum L. (G‐V) were investigated using relative quantitative method of electrospray ionization mass spectrometry (ESI‐MS) and UPLC‐MSⁿ, respectively. The relative contents of most ginsenosides were reduced in the extract of G‐V. Furthermore, ginsenosides Rb₁, Rb₂, Rc and Rd could be metabolized to Rd, F2 and C‐K in rat intestinal bacteria. The metabolic speeds of Rb₁, Rb₂ and Rc in the G‐V extracts at ratios of 10:5, 10:7 and 10:10 and the metabolic rates of ginsenosides Rb₁, Rb₂ and Rc to Rd, Rd to F2 in all compatibility extracts were lower than that in the P. ginseng extract. In conclusion, this study illustrated the mechanism of effect‐reducing by comparison of the relative contents and metabolic profiles of ginsenosides after compatibility of P. ginseng and V. nigrum L.     

Simplified ultrasonically- and microwave-assisted solvent extractions for the determination of ginsenosides in powdered Panax ginseng rhizomes using liquid chromatography with UV absorbance or electrospray mass spectrometric detection

New approaches for the recovery of ginsenosides are presented that greatly simplify the liquid chromatographic (LC) determination of the total content of eight ginsenosides (Rb₁, Rb₂, Rc, Rd, Re, Rf, Rg₁ and Rg₂) in powdered Panax ginseng rhizomes. The extraction protocols not only recover the neutral ginsenosides, but also simultaneously incorporate base-catalyzed hydrolysis of the malonyl-ginsenosides using dilute potassium hydroxide added to the methanol–water extractant. This eliminates the need for an independent extraction step followed by acid- or base-catalyzed hydrolysis. Both ultrasonically-assisted and microwave-assisted extraction methods are developed. The optimization of these simplified methods to remove pendant malonate esters, while retaining the glycosidic linkages, was determined by LC through variation of the extraction/hydrolysis time, order of hydrolysis reagent addition, and evaluation of multiple extractions. A comparison of the ginsenoside profiles obtained with and without addition of base to the extractant solution was made using LCMS with positive-mode electrospray ionization (ESI⁺) detection. A number of malonyl-ginsenosides were tentatively identified by their mass spectral fragmentation spectra and indicating that they were converted to the free ginsenosides by the new extraction/hydrolysis procedure.

Major ginsenoside contents in rhizomes of Panax sokpayensis and Panax bipinnatifidus

This study compared eight major ginsenosides (Rg1, Rg2, Rf, Re, Rd, Rc, Rb1 and Rb2) between Panax sokpayensis and Panax bipinnatifidus collected from Sikkim Himalaya, India. High-performance liquid chromatographic analysis revealed that all major ginsenosides were present in the rhizomes of P. sokpayensis except ginsenoside Rc, whereas ginsenoside Rf, Rc and Rb2 were not detected in P. bipinnatifidus.     

177 Saponins,Including 11 New Compounds in Wild Ginseng Tentatively Identified via HPLC-IT-TOF-MSn,and Differences among Wild Ginseng,Ginseng under Forest,and Cultivated Ginseng

Wild ginseng (W-GS), ginseng under forest (F-GS, planted in mountain forest and growing in natural environment), and cultivated ginseng (C-GS) were compared via HPLC-DAD and HPLC-IT-TOF-MSⁿ. A total of 199 saponins, including 16 potential new compounds, were tentatively identified from 100 mg W-GS (177 saponins in W-GS with 11 new compounds), F-GS (56 saponins with 1 new compound), and C-GS (60 saponins with 6 new compounds). There were 21 saponins detected from all the W-GS, F-GS, and C-GS. Fifty saponins were only detected from W-GS, including 23 saponins found in ginseng for the first time. Contents of ginsenosides Re (12.36–13.91 mg/g), Rh₁ (7.46–7.65 mg/g), Rd (12.94–12.98 mg/g), and the total contents (50.52–55.51 mg/g) of Rg₁, Re, Rf, Rb₁, Rg₂, Rh₁, and Rd in W-GS were remarkably higher than those in F-GS (Re 1.22–3.50 mg/g, Rh₁ 0.15–1.49 mg/g, Rd 0.19–1.49 mg/g, total 5.69–18.74 mg/g), and C-GS (Re 0.30–3.45 mg/g, Rh₁ 0.05–3.42 mg/g, Rd 0.17–1.68 mg/g, total 2.99–19.55 mg/g). Contents of Re and Rf were significantly higher in F-GS than those in C-GS (p < 0.05). Using the contents of Re, Rf, or Rb₁, approximately a half number of cultivated ginseng samples could be identified from ginseng under forest. Contents of Rg₁, Re, Rg₂, Rh₁, as well as the total contents of the seven ginsenosides were highest in ginseng older than 15 years, middle–high in ginseng between 10 to 15 years old, and lowest in ginseng younger than 10 years. Contents of Rg₁, Re, Rf, Rb₁, Rg₂, and the total of seven ginsenosides were significantly related to the growing ages of ginseng (p < 0.10). Similarities of chromatographic fingerprints to W-GS were significantly higher (p < 0.05) for F-GS (median: 0.824) than C-GS (median: 0.745). A characteristic peak pattern in fingerprint was also discovered for distinguishing three types of ginseng. Conclusively, wild ginseng was remarkably superior to ginseng under forest and cultivated ginseng, with ginseng under forest slightly closer to wild ginseng than cultivated ginseng. The differences among wild ginseng, ginseng under forest, and cultivated ginseng in saponin compositions and contents of ginsenosides were mainly attributed to their growing ages.     

人参皂甙的反相高效液相色谱多台阶梯度优化方法

建立了一种反相高效液相色谱多台阶梯度分离人参皂甙的方法.该方法以乙腈-水溶液为流动相,通过一系列等度实验,获得了8种人参皂甙Rg1,Re,Rf,Rg2,Rb1,Rc,Rb2和Rd的色谱保留参数,发现两参数保留方程不适合用于人参皂甙这种天然产物的分离条件的优化,而三参数保留方程的高精度才可满足预测的要求.在三参数保留方程的基础上,通过计算确定了8种人参皂甙(包括3台阶梯度)的液相色谱分离条件.通过实验对此优化条件进行了验证,实验结果显示了较好的预测精度和分离度.将本方法用于分离人参皂甙,分析时间短且分离度高,显示了等度台阶梯度优化方法对确定色谱分离条件的优越性.     

Metabolite profiling of ginsenosides in rat plasma,urine and feces by LC–MS/MS and its application to a pharmacokinetic study after oral administration of Panax ginseng extract

Panax ginseng is widely consumed as a functional food in the form of tea, powder, capsules, among others, and possesses a range of pharmacological activities including adaptogenic, immune‐modulatory, anti‐tumor, anti‐aging and anti‐inflammatory effects. The aim of this study was to identify and quantify the major ginsenosides and their metabolites in rat plasma, urine and feces after administration of P. ginseng extract using LC–MS/MS. We collected rat plasma samples at 0.5, 1, 2, 4, 8, 12, 24 and 48 h, and the amounts of urine and fecal samples accumulated in 24 h. Fourteen major ginsenosides and their metabolites were observed in fecal samples at high levels; however, low levels of 11 ginsenosides were detected in urine samples. The pharmacokinetics of the major ginsenosides and their metabolites was investigated in plasma. The results indicated that the maximum plasma concentration, time to maximum concentration and area under the curve of compound K were significantly greater than those of other ginsenosides. This study thus provides valuable information for drug development and clinical application of P. ginseng.     

Determination of marker constituents in radix Glycyrrhizae and radix Notoginseng by near infrared spectroscopy

High-performance liquid chromatographic (HPLC) methods were developed for the determination of glycyrrhizin in radix Glycyrrhizae and ginsenosides Rb₁, Rb₂, Rc, Rd, Re, Rf and Rg₁ in radix Notoginseng. These methods were used as reference methods for near-infrared (NIR) spectroscopy. Spectroscopic calibrations were developed for the determination of glycyrrhizin, the total content of ginsenosides and the individual major ginsenosides Rb₁, Rd, Re and Rg₁. Standard errors of cross validation (SECV) were 1.22 mg g^–1 for glycyrrhizin (concentration range 21.3–34.1 mg g^–1) and 0.99 mg g^–1 for the sum of ginsenosides (concentration range 55.3–¶71.1 mg g^–1). The corresponding coefficients of determination (R²) were 0.94 and 0.98, respectively. The SECVs were generally less than a factor of 2.5 of the repeatability standard deviation of the HPLC methods.     

Comprehensive Investigation on Ginsenosides in Different Parts of a Garden-Cultivated Ginseng Root and Rhizome

Background: Ginseng is widely used as herb or food. Different parts of ginseng have diverse usages. However, the comprehensive analysis on the ginsenosides in different parts of ginseng root is scarce. Methods: An ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF/MS) combined with UNIFI informatics platform and ultra-high-performance liquid chromatography-charged aerosol detection (UHPLC-CAD) were employed to evaluate the different parts of cultivated ginseng root. Results: 105 ginsenosides including 16 new compounds were identified or tentatively characterized. 22 potential chemical markers were identified, 20, 17, and 19 for main root (MR) and fibrous root (FR), main root (MR) and branch root (BR), and main root (MR) and rhizome (RH), respectively. The relative contents of Re, Rb₁, 20(R)-Rh₁, Rd, and Rf were highest in FR. The relative content of Rg₁ was highest in RH. The total relative content of pharmacopoeia indicators Rg₁, Re, and Rb₁ was highest in FR. Conclusion: The differences among these parts were the compositions and relative contents of ginsenosides. Under our research conditions, the peak area ratio of Rg₁ and Re could distinguish the MR and FR samples. Fibrous roots showed rich ingredients and high ginsenosides contents which should be further utilized.     

Development of Green and Efficient Extraction of Bioactive Ginsenosides from Panax ginseng with Deep Eutectic Solvents

The extraction of active constituents from natural sources in a green and efficient manner is considered an important field in the pharmaceutical industry. In recent years, deep eutectic solvents (DESs), a new type of green solvent, have attracted increasing attention. Therefore, we aimed to establish a green and high-efficiency extraction method for ginsenosides based on DESs. This study takes Panax ginseng as a model sample. Eighteen different DESs were produced to extract polar ginsenosides. Ultrasound-assisted extraction (UAE) was applied for simplicity and efficiency. A binary DES synthesized using choline chloride and urea at a proportion of 1:2 prepared by a heating stirring method is proven to be more effective than other solvents, such as the widely used 70% ethanol for the extraction of ginsenosides. Three variables that might affect the extraction, including the DES content in the extraction solvent, liquid/solid ratio, and ultrasound extraction time, were evaluated for optimization. The optimum extraction conditions for ginsenosides were determined as follows: DES water content of 20 wt%, liquid/solid ratio of 15 mL g⁻¹, and an ultrasonic extraction time of 15 min. The extraction yield for the optimized method is found to be 31% higher than that for 70% ethanol, which achieves efficient extraction. This study shows that DESs are available to extract ginsenosides for use in traditional Chinese medicine. The discovery also contributes to further research into the green extraction of ginsenosides.     

Rapid differentiation of Panax ginseng and Panax quinquefolius by matrix-assisted laser desorption/ionization mass spectrometry

A matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS)-based method has been developed for rapid differentiation between Panax ginseng and Panax quinquefolius, two herbal medicines with similar chemical and physical properties but different therapeutic effects. This method required only a small quantity of samples, and the herbal medicines were analyzed by MALDI-MS either after a brief extraction step, or directly on the powder form or small pieces of raw samples. The acquired MALDI-MS spectra showed different patterns of ginsenosides and small chemical molecules between P. ginseng and P. quinquefolius, thus allowing unambiguous differentiation between the two Panax species based on the specific ions, intensity ratios of characteristic ions or principal component analysis. The approach could also be used to differentiate red ginseng or P. quinquefolius adulterated with P. ginseng from pure P. ginseng and pure Panax quinquefolium. The intensity ratios of characteristic ions in the MALDI-MS spectra showed high reproducibility and enabled quantitative determination of ginsenosides in the herbal samples and percentage of P. quinquefolius in the adulterated binary mixture. The method is simple, rapid, robust, and can be extended for analysis of other herbal medicines.     

人参皂苷的仿生提取研究

通过体外模拟胃肠道环境,建立一种提取人参皂苷的仿生方法.考察了提取条件对配制的仿生胃液和仿生肠液提取人参皂苷浓度的影响.基于高效液相色谱-三重四极杆质谱的多反应监测模式建立定量分析Re,Rg1,20(S)-Rf,Rb1,Ro,Rc,Rb2,Rd等8种人参皂苷的方法,并比较了仿生和超声两种提取方法的人参皂苷提取效率.结果...     

Isolation and Purification of Ginsenosides from Plant Extract of Panax quinquefolium L. by High Performance Centrifugal Partition Chromatography Coupled with ELSD

A high performance centrifugal partition chromatography (HPCPC) combined with evaporative light scattering detection (ELSD) was developed for the separation and purification of ginsenosides from Panax quinquefolium. Three compounds, ginsenosides Rc, Rb₁, and Re were isolated and purified by HPCPC using an optimized two-phase solvent system composed of ethyl acetate–n-butanol–water (1:1:2, v/v/v). The purities of the three ginsenosides were 96.5, 97.6, and 98.5%, respectively as determined by liquid chromatography (LC–ELSD). The CPC fractions were analyzed by LC–ELSD and electrospray ion source mass spectroscopy (ESI-MSⁿ) in negative ion mode. The identification of the ginsenosides Rc, Rb₁, and Re in the extract of P. quinquefolium was based on matching their retention times, the detection of the molecular ions, and the fragment ions of the molecular ion obtained in the CID experiments with those of the authentic standards and data reported in the literature. The results demonstrate that HPCPC coupled with ELSD is a feasible and efficient technique for systematic isolation of non-chromophoric components from traditional medicinal herbs.     

Chemical and Pharmacological Profiling of Wrightia coccinea (Roxb. Ex Hornem.) Sims Focusing Antioxidant,Cytotoxic, Antidiarrheal,Hypoglycemic, and Analgesic Properties

The aim of the study was to conduct phytochemical and pharmacological investigations of Wrightia coccinea (Roxb. ex Hornem.) Sims via several in vitro, in vivo, and in silico models. A total of four compounds were identified and isolated from the methanol extract of the bark and the methanol extract of the seed pulp of W. coccinea through successive chromatographic techniques and were characterized as 3β-acetyloxy-olean-12-en-28-ol (1), wrightiadione (2), 22β-hydroxylupeol (3), and β-sitosterol (4) by spectroscopic analysis. The aqueous fraction of the bark and chloroform fraction of the fruits provided the most potent antioxidant capacity (IC₅₀ = 7.22 and 4.5 µg/mL, respectively) in DPPH free radical scavenging assay compared with the standard ascorbic acid (IC₅₀ = 17.45 µg/mL). The methanol bark extract and the methanol fruit coat extract exerted anti-diarrheal activity by inhibiting 74.55 ± 0.67% and 77.78 ± 1.5% (mean ± SEM) of the diarrheal episode in mice, respectively, after four hours of loading the samples. In the hypoglycemic test, the methanol bark extract and the methanol fruit coat extract (400 mg/kg) produced a significant (p < 0.05) reduction in the blood glucose level in mice. Both doses of the plant extracts (200 mg/kg and 400 mg/kg) used in the study induced a significant (p < 0.05) increase in pain reaction time. The in vitro and in vivo findings were supported by the computational studies. The isolated compounds exhibited higher binding affinity compared with the standard drugs towards the active binding sites of glutathione reductase, epidermal growth factor receptor (EGFR), kappa opioid receptor, glucose transporter 3 (GLUT 3), Mu opioid receptor, and cyclooxygenase 2 (COX-2) proteins due to their potent antioxidant, cytotoxic, anti-diarrheal, hypoglycemic, and central and peripheral analgesic properties, respectively. The current findings concluded that W. coccinea might be a potential natural source for managing oxidative stress, diarrhea, hyperglycemia, and pain. Further studies are warranted for extensively phytochemical screening and establishing exact mechanisms of action.     

Retention behavior of ginsenosides on a poly(vinyl alcohol)-bonded stationary phase in hydrophilic interaction chromatography

The influences of the organic component of the mobile phase and the column temperature on the retention of ginsenosides on a poly(vinyl alcohol) (PVA) bonded stationary phase operated under hydrophilic interaction chromatographic mode were investigated. The retention of the ginsenosides was found to increase with increasing amount of acetonitrile (MeCN) in the mobile phase, which is typical of hydrophilic interaction chromatographic behavior. It was also found that the retention of the analytes was highly affected by the type of the organic modifier used. Aqueous MeCN (75–90%) gave the most satisfactory retention and separation of ginsenosides Rf, Rg1, Rd, Re, Rc, Rb2 and Rb1 compared with aqueous methanol, isopropyl alcohol or tetrahydrofuran at the same composition levels. The effects of the different types of organic modifiers on the retention of the analytes were attributed to their solvent strength and hydrogen-bond accepting/donating properties. The effect of temperature on the retention of ginsenoside on the PVA-bonded phase was assessed by constructing van’t Hoff plots for two temperature ranges: subambient (273–293 K) and ambient-elevated (298–333 K) temperatures. van’t Hoff plots for all analytes were linear at the two temperature intervals; however, the slopes of the lines corresponding to ginsenosides Rg1 and Re were completely different from those for the rest of the analytes especially in the subambient temperature range. Enthalpy-entropy compensation (EEC) studies were conducted to verify the difference in thermodynamics observed for ginsenosides Rg1 and Re compared with the other analytes. EEC plots showed that Rf, Rd, Rc, Rb2 and Rb1 were possibly retained by the same retention mechanism, which was completely different from that of Rg1 and Re at subambient temperatures. Retention prediction models were derived using multiple linear regression to identify solute attributes that affected the retention of the analytes on the PVA-bonded phase. The mathematical models derived revealed that the number of hydrogen-bond donors and the ovality of the molecules are important molecular properties that govern the retention of the compounds on the chromatographic system.     

The use of linear ion trap for qualitative analysis of phytochemicals in Korean ginseng tea

A new approach to qualitative analysis of ginsenosides in challenging matrices was developed on the basis of high‐performance liquid chromatography/tandem mass spectrometry. Using the extracts from samples of ginseng tea, the approach was validated. Analysis of extracts was carried out using a reversed‐phase chromatography with SB‐C₁₈ sorbent. For compound identification, electrospray ionization and a quadrupole/linear ion trap mass‐spectrometer in different modes were used. A meticulous study of the fragmentation of ginsenosides in the linear ion trap and its application for analysis of these compounds was performed in this work. The accuracy of the identification was proven with standards of ginsenosides Rb1, Rg1, Re, Rf, Rd, Rb2, Rb3 and Rc. Copyright © 2012 John Wiley & Sons, Ltd.     

Adsorption and desorption characteristics of ginsenosides from Panax ginseng C. A. Meyer on middle‐pressure chromatogram isolated gels

Four types of middle‐pressure chromatogram isolated gels are evaluated for adsorption or desorption characteristics of ginsenosides from Panax ginseng. Among them, SP207SS and SP2MGS were selected for dynamic investigations based on their static adsorption or desorption capacity of total ginsenoside. Their adsorption kinetics was better explained by pseudosecond‐order model and isotherms were preferably fitted to Langmuir model. Dynamic breakthrough experiments indicated an optimum sample loading speed of 4 bed volume/h for either SP207SS or SP2MGS. Desorption speed was determined to be 2 bed volume/h according to desorption amount of total ginsenoside in their effluents. Eight ginsenosides were identified and quantified by high performance liquid chromatography‐triple quadropole‐mass spectrometry in total ginsenoside extract and different fractions during stepwise dynamic elution. For SP207SS, 27.62% of loaded ginsenosides was detected in 40% ethanol fraction, while 59.12% of them were found in 60% ethanol fraction. As on SP2MGS, the number went to 53.71 and 44.43%, respectively. Recovery rate of ginsenosides were calculated to 78.65% for SP207SS and 89.53% for SP2MGS, respectively. Intriguingly, content of Rg1 and Re in 40% ethanol fraction from SP207SS became 20.1 and 18.6 times higher than that in total ginsenoside extract by one‐step elution, which could be leveraged for the facile enrichment of these two ginsenosides from natural sources.     

A quantified ginseng (Panax ginseng C.A. Meyer) extract influences lipid acquisition and increases adiponectin expression in 3T3-L1 cells

A Panax ginseng extract (PGE) with a quantified amount of ginsenosides was utilized to investigate its potential to inhibit proliferation, influence lipid acquisition and adiponectin expression in 3T3-L1 cells. Seven fingerprint ginsenosides were quantified using high performance liquid chromatography and their respective molecular weights were further confirmed via LC-ESI-MS analysis from four different extraction methods. Extraction using methanol under reflux produced significantly higher amounts of ginsenosides. The methanol extract consisted of Rg1 (47.40 ± 4.28 mg/g, dry weight of extract), Re (61.62 ± 5.10 mg/g), Rf (6.14 ± 0.28 mg/g), Rb1 (21.73 ± 1.29 mg/g), Rc (78.79 ± 4.15 mg/g), Rb2 (56.80 ± 3.79 mg/g), Rd (5.90 ± 0.41 mg/g). MTT analysis showed that PGE had a concentration-dependent cytotoxic effect on 3T3-L1 preadipocyte and the LC(50) value was calculated to be 18.2 ± 5 μg/mL. Cell cycle analysis showed minimal changes in all four phases. Differentiating adipocytes treated with ginseng extract had a visible decrease in lipid droplets formation measured by Oil red O staining. Consequently, triglycerides levels in media significantly (P < 0.05) decreased by 39.5% and 46.1% when treated at concentrations of 1 μg/mL and 10 μg/mL compared to untreated control cells. Western blot analysis showed that the adiponectin protein expression was significantly (P < 0.05) increased at 10 μg/mL, but not at 1 μg/mL. A quantified PGE reduced the growth of 3T3-L1 cells, down-regulated lipid accumulation and up-regulated adiponectin expression in the 3T3-L1 adipocyte cell model.     

High Performance Liquid Chromatographic Separation and Quantitative Analysis of Ginsenosides Using a Polyvinyl Alcohol-Bonded Stationary Phase

A simple high performance liquid chromatographic assay for the simultaneous quantitative analysis of seven ginsenosides, Rb1, Rb2, Rc, Rd, Re, Rf and Rg1 in commercial ginseng products is described. Chromatographic separation of the analytes was achieved in less than 20 min using a polyvinyl alcohol-bonded column with UV detection at 203 nm. Optimization of chromatographic conditions was determined by a three-factor central composite design, the variables being the percentage of acetonitrile in the mobile phase, column temperature and flow rate. A full quadratic model was found to be adequate in describing the separation of ginsenosides on the polyvinyl alcohol-bonded stationary phase. Complete separation of seven ginsenosides was achieved using acetonitrile–water (82.5/17.5) as the mobile phase run isocratically at a flow rate of 298 μL min^?1 and with the column temperature at 9 °C. The developed method was validated over the range of 10–120 μg mL^?1 using a 5 μL sample injection volume. Intra- and inter-day variation for three ginsenoside standards (Rf, Rd and Rb1) at three concentration levels ranged from 0.07 to 0.83% expressed as the relative standard deviation. The accuracy based on the nominal concentration values at three concentration levels was in the range 98.7–100.8%. The limit of detection was between 0.43 and 1.03 μg mL^?1 while the limit of quantification was from 1.42 to 3.13 μg mL^?1. The method is found to be applicable for the determination of ginsenosides in commercial ginseng products.