Evaluation of the Saponin Content in Panax vietnamensis Acclimatized to Lam Dong Province by HPLC-UV/CAD.

Panax vietnamensis, or Vietnamese ginseng (VG), an endemic Panax species in Vietnam, possesses a unique saponin profile and interesting biological activities. This plant is presently in danger of extinction due to over-exploitation, resulting in many preservation efforts towards the geographical acclimatization of VG. Yet, no information on the saponin content of the acclimatized VG, an important quality indicator, is available. Here, we analyzed the saponin content in the underground parts of two- to five-year-old VG plants acclimatized to Lam Dong province. Nine characteristic saponins, including notoginsenoside-R1, ginsenoside-Rg1, -Rb1, -Rd, majonoside-R1, -R2 vina-ginsenoside-R2, -R11, and pseudoginsenoside-RT4, were simultaneously determined by HPLC coupled with UV and with a charged aerosol detector (CAD). Analyzing the results illustrated that the detection of characteristic ocotillol-type saponins in VG by CAD presented a superior capacity compared with that of UV, thus implying a preferential choice of CAD for the analysis of VG. The quantitative results indicating the saponin content in the underground parts of VG showed an increasing tendency from two to five years old, with the root and the rhizome exhibiting different saponin accumulation patterns. This is the first study that reveals the preliminary success of VG acclimatization and thereby encourages the continuing efforts to develop this valuable saponin-rich plant.

Characterization and Antioxidant Activity Determination of Neutral and Acidic Polysaccharides from Panax Ginseng C. A. Meyer.

Panax ginseng (P. ginseng) is the most widely consumed herbal plant in Asia and is well-known for its various pharmacological properties. Many studies have been devoted to this natural product. However, polysaccharide's components of ginseng and their biological effects have not been widely studied. In this study, white ginseng neutral polysaccharide (WGNP) and white ginseng acidic polysaccharide (WGAP) fractions were purified from P. ginseng roots. The chemical properties of WGNP and WGAP were investigated using various chromatography and spectroscopy techniques, including high-performance gel permeation chromatography, Fourier-transform infrared spectroscopy, and high-performance liquid chromatography with an ultra-violet detector. The antioxidant, anti-radical, and hydrogen peroxide scavenging activities were evaluated in vitro and in vivo using Caenorhabditis elegans as the model organism. Our in vitro data by ABTS (2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid), reducing power, ferrous ion chelating, and hydroxyl radical scavenging activity suggested that the WGAP with significantly higher uronic acid content and higher molecular weight exhibits a much stronger antioxidant effect as compared to that of WGNP. Similar antioxidant activity of WGAP was also confirmed in vivo by evaluating internal reactive oxygen species (ROS) concentration and lipid peroxidation. In conclusion, WGAP may be used as a natural antioxidant with potent scavenging and metal chelation properties.

Accumulation of Saponins in Underground Parts of Panax vietnamensis at Different Ages Analyzed by HPLC-UV/ELSD.

Panax vietnamensis (PV), a wild Panax species discovered in Vietnam in 1973, has been increasingly overexploited due to its economic value and therapeutic uses. This resulted in the development of PV cultivation to meet the market demand. There is little information on the accumulation of saponins in PV during cultivation, but this information could serve as an indication of the appropriate harvest time. In this study we developed an HPLC-UV/ELSD method to simultaneously determine the content of 10 characteristic saponins in PV from 2-7 years old, including G-Rb1, G-Rd, G-Rg1, G-Re, N-R1, M-R1, M-R2, V-R2, V-R11, and p-RT4. The result indicated that from 2 to 5 years, the content of saponins in PV rhizome and radix increase 3.02 and 4.2 times, respectively, whereas from 5 to 7 years, no significant changes were observed. Hence, our study suggests that after 5 years of growth could be considered as an appropriate time for PV to be harvested. Among the analyzed saponins, G-Rg1, G-Rb1, G-Rd, and especially M-R2 were the major saponins that contributed to the change of PV's saponin content through the years. In addition, the developed and validated HPLC method was proven to be reliable and effective for quality control of PV.

Adiponectin-Secretion-Promoting Phenylethylchromones from the Agarwood of Aquilaria malaccensis.

The therapeutic potential of adiponectin regulation has received interest because of its association with diverse human disease conditions, such as diabetes, obesity, atherosclerosis, and cancer. Phenylethylchromone derivatives from Aquilaria malaccensis-derived agarwood promoted adiponectin secretion during adipogenesis in human bone marrow mesenchymal stem cells, and 5,6-dihydroxy-2-(2-phenylethyl)chromone (1) was identified as a new chromone derivative. A target identification study with the most potent adiponectin-secretion-promoting phenylethylchromones, 6-methoxy-2-(2-phenylethyl)chromone (3) and 7-methoxy-2-(2-phenylethyl)chromone (4), showed that they are PPARγ partial agonists. Therefore, the diverse therapeutic effects of agarwood are associated with a PPARγ-mediated adiponectin-secretion-promoting mechanism.

High-Throughput Omics and Statistical Learning Integration for the Discovery and Validation of Novel Diagnostic Signatures in Colorectal Cancer.

The advancement of bioinformatics and machine learning has facilitated the discovery and validation of omics-based biomarkers. This study employed a novel approach combining multi-platform transcriptomics and cutting-edge algorithms to introduce novel signatures for accurate diagnosis of colorectal cancer (CRC). Different random forests (RF)-based feature selection methods including the area under the curve (AUC)-RF, Boruta, and Vita were used and the diagnostic performance of the proposed biosignatures was benchmarked using RF, logistic regression, naïve Bayes, and k-nearest neighbors models. All models showed satisfactory performance in which RF appeared to be the best. For instance, regarding the RF model, the following were observed: mean accuracy 0.998 (standard deviation (SD) < 0.003), mean specificity 0.999 (SD < 0.003), and mean sensitivity 0.998 (SD < 0.004). Moreover, proposed biomarker signatures were highly associated with multifaceted hallmarks in cancer. Some biomarkers were found to be enriched in epithelial cell signaling in Helicobacter pylori infection and inflammatory processes. The overexpression of TGFBI and S100A2 was associated with poor disease-free survival while the down-regulation of NR5A2, SLC4A4, and CD177 was linked to worse overall survival of the patients. In conclusion, novel transcriptome signatures to improve the diagnostic accuracy in CRC are introduced for further validations in various clinical settings.

Increase in Protective Effect of Panax vietnamensis by Heat Processing on Cisplatin-Induced Kidney Cell Toxicity.

Cisplatin is a platinum-based anticancer agent used for treating a wide range of solid cancers. One of the side effects of this drug is its severe nephrotoxicity, limiting the safe dose of cisplatin. Therefore, many natural products have been studied and applied to attenuate the toxicity of this compound. In this study, we found that steamed Vietnamese ginseng (Panax vietnamensis) could significantly reduce the kidney damage of cisplatin in an in vitro model using porcine proximal tubular LLC-PK1 kidney cells. From processed ginseng under optimized conditions (120 °C, 12 h), we isolated seven compounds (20(R,S)-ginsenoside Rh2, 20(R,S)-ginsenoside Rg3, ginsenoside Rk1, ginsenoside-Rg5, and ocotillol genin) that showed kidney-protective potential against cisplatin toxicity. By comparing the 50% recovery concentration (RC50), the R form of ginsenoside, Rh2 and Rg3, had RC50 values of 6.67 ± 0.42 µM and 8.39 ± 0.3 µM, respectively, while the S forms of ginsenoside, Rh2 and Rg3, and Rk1, had weaker protective effects, with RC50 ranging from 46.15 to 88.4 µM. G-Rg5 and ocotillol, the typical saponin of Vietnamese ginseng, had the highest RC50 (180.83 ± 33.27; 226.19 ± 66.16, respectively). Our results suggest that processed Vietnamese gingseng (PVG), as well as those compounds, has the potential to improve kidney damage due to cisplatin toxicity.

Integrated metabolomics signature for assessing the longevity of Panax ginseng seeds.

BACKGROUND: Panax ginseng seeds have strong dormancy and a prolonged germination period in comparison to other seeds; thus, it is a great challenge to propagate ginseng. Seed longevity is closely associated with germination rate and viability, so we assumed that if a seed loses its viability, specific metabolic alterations regarding plant growth factors might occur. In this study, we divided ginseng seeds into normal and accelerated-aging groups. Both groups were treated with gibberellic acid, which is one of the most important plant-growth regulators. Afterward, gas chromatography-mass spectrometry (GC-MS) was used to analyze the samples, to identify the metabolic alterations between the two groups. RESULTS: Forty-four endogenous metabolites in normal and accelerated aging groups were putatively identified. To determine the differential significance of these metabolites, t-tests and fold-change analysis were conducted followed by principal component analysis and partial least-squares discriminant analysis to determine the metabolites that showed distinct responses between the groups. Among the differentially expressed metabolites (P value < 0.05 and FDR < 0.1), nine metabolites were selected as potential biomarker candidates for the prediction of seed longevity. CONCLUSION: Nine metabolites related to ginseng seed longevity were identified by comparing metabolomes. Our findings suggest that ginseng propagation can be facilitated by the regulation of these distinctive metabolic features of the seeds. © 2019 Society of Chemical Industry.

Aquilanols A and B, Macrocyclic Humulene-Type Sesquiterpenoids from the Agarwood of Aquilaria malaccensis.

Four new and five known sesquiterpenoids were isolated from the agarwood of Aquilaria malaccensis. Aquilanols A and B (1 and 2) have an unprecedented macrocyclic humulene structure with a bicyclic 7/10 ring system. Compound 2 was obtained as a scalemic mixture that was resolved by HPLC analysis using a chiral column. Their structures were deduced based on spectroscopic data analysis, and the absolute configurations were unambiguously determined by X-ray crystallographic data and ECD spectroscopic analysis. A putative biosynthetic pathway of these sesquiterpenoids is proposed.

Solid-phase extraction assisted dispersive liquid-liquid microextraction based on solidification of floating organic droplet to determine sildenafil and its analogues in dietary supplements.

A novel analytical method for the simultaneous determination of the concentration of sildenafil and its five analogues in dietary supplements using solid-phase extraction assisted reversed-phase dispersive liquid-liquid microextraction based on solidification of floating organic droplet combined with ion-pairing liquid chromatography with an ultraviolet detector was developed. Parameters that affect extraction efficiency were systematically investigated, including the type of solid-phase extraction cartridge, pH of the extraction environment, and the type and volume of extraction and dispersive solvent. The method linearity was in the range of 5.0-100 ng/mL for sildenafil, homosildenafil, udenafil, benzylsildenafil, and thiosildenafil and 10-100 ng/mL for acetildenafil. The coefficients of determination were ≥0.996 for all regression curves. The sensitivity values expressed as limit of detection were between 2.5 and 7.5 ng/mL. Furthermore, intraday and interday precisions expressed as relative standard deviations were less than 5.7 and 9.9%, respectively. The proposed method was successfully applied to the analysis of sildenafil and its five analogues in complex dietary supplements.

Stereoisomer-specific anticancer activities of ginsenoside Rg3 and Rh2 in HepG2 cells: disparity in cytotoxicity and autophagy-inducing effects due to 20(S)-epimers.

Autophagy has been an emerging field in the treatment of hepatic carcinoma since anticancer therapies were shown to ignite autophagy in vitro and in vivo. Here we report that ginsenoside Rg3 and Rh2, major components of red ginseng, induce apoptotic cell death in a stereoisomer-specific fashion. The 20(S)-forms of Rg3 and Rh2, but not their respective 20(R)-forms, promoted cell death in a dose-dependent manner accompanied by downregulation of Bcl2 and upregulation of Fas, resulting in apoptosis of HepG2 cells with poly ADP ribose polymerase cleavage. The LD50 value [45 µM for Rg3(S), less than 10 µM for Rh2(S)] and gross morphological electron microscopic observation revealed more severe cellular damage in cells treated with Rh2(S) than in those treated with Rg3(S). Both Rg3(S) and Rh2(S) also induced autophagy when undergoing induced apoptosis. Inhibition of autophagy with lysosomotrophic agents significantly potentiated the cellular damage, implying a favorable switch of the cell fate to tumor cell death. Blocking intracellular calcium with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis(acetoxymethyl ester) (BAPTA-AM) restored the cell death induced by both Rg3(S) and Rh2(S). Our results suggest that the 20(S)-forms of Rg3 and Rh2 in red ginseng possess more potent antitumor activity with autophagy than their 20(R)-forms via calcium-dependent apoptosis.

Ginseng Saponins in Different Parts of Panax vietnamensis.

Chemical and pharmacological studies of Panax vietnamensis (Vietnamese ginseng; VG) have been reported since its discovery in 1973. However, the content of each saponin in different parts of VG has not been reported. In this study, 17 ginsenosides in the different underground parts of P. vietnamensis were analyzed by HPLC/evaporative light scattering detector (ELSD). Their contents in the dried rhizome, radix, and fine roots were 195, 156, and 139 mg/g, respectively, which were extremely high compared to other Panax species. The content of protopanaxatriol (PPT)-type saponins were not much different among underground parts; however, the content of protopanaxadiol (PPD)- and ocotillol (OCT)-type saponins were greatly different. It is noteworthy that the ginsenoside pattern in the fine roots is different from other underground parts. In particular, despite the content of PPD-type saponins being the highest in the fine roots, which is similar to other Panax species, the total content of saponins was the lowest in the fine roots, which is different from other Panax species. The ratios of PPT : PPD : OCT-type saponins were 1 : 1.7 : 7.8, 1 : 1.6 : 5.5, and 1 : 4.8 : 3.3 for the rhizome, radix, and fine roots, respectively. OCT-type saponins accounted for 36-75% of total saponins and contributed mostly to the difference in the total saponin content of each part.

Novel approach for analysis of bronchoalveolar lavage fluid (BALF) using HPLC-QTOF-MS-based lipidomics: lipid levels in asthmatics and corticosteroid-treated asthmatic patients.

To better understand the respiratory lipid phenotypes of asthma, we developed a novel method for lipid profiling of bronchoalveolar lavage fluid (BALF) using HPLC-QTOF-MS with an internal spectral library and high-throughput lipid-identifying software. The method was applied to BALF from 38 asthmatic patients (18 patients with nonsteroid treated bronchial asthma [NSBA] and 20 patients with steroid treated bronchial asthma [SBA]) and 13 healthy subjects (NC). We identified 69 lipids, which were categorized into one of six lipid classes: lysophosphatidylcholine (LPC), phosphatidylcholine (PC), phosphatidylglycerol (PG), phosphatidylserine (PS), sphingomyelin (SM) and triglyceride (TG). Compared with the NC group, the individual quantity levels of the six classes of lipids were significantly higher in the NSBA subjects. In the SBA subjects, the PC, PG, PS, SM, and TG levels were similar to the levels observed in the NC group. Using differentially expressed lipid species (p value < 0.05, FDR < 0.1 and VIP score of PLS-DA > 1), 34 lipid biomarker candidates with high prediction performance between asthmatics and controls were identified (AUROC > 0.9). These novel findings revealed specific characteristics of lipid phenotypes in asthmatic patients and suggested the importance of future research on the relationship between lipid levels and asthma.

Modulation of lipid kinase PI4KIIα activity and lipid raft association of presenilin 1 underlies γ-secretase inhibition by ginsenoside (20S)-Rg3.

Amyloid ß-peptide (Aß) pathology is an invariant feature of Alzheimer disease, preceding any detectable clinical symptoms by more than a decade. To this end, we seek to identify agents that can reduce Aß levels in the brain via novel mechanisms. We found that (20S)-Rg3, a triterpene natural compound known as ginsenoside, reduced Aß levels in cultured primary neurons and in the brains of a mouse model of Alzheimer disease. The (20S)-Rg3 treatment induced a decrease in the association of presenilin 1 (PS1) fragments with lipid rafts where catalytic components of the γ-secretase complex are enriched. The Aß-lowering activity of (20S)-Rg3 directly correlated with increased activity of phosphatidylinositol 4-kinase IIα (PI4KIIα), a lipid kinase that mediates the rate-limiting step in phosphatidylinositol 4,5-bisphosphate synthesis. PI4KIIα overexpression recapitulated the effects of (20S)-Rg3, whereas reduced expression of PI4KIIα abolished the Aß-reducing activity of (20S)-Rg3 in neurons. Our results substantiate an important role for PI4KIIα and phosphoinositide modulation in γ-secretase activity and Aß biogenesis.

Combination of a sub-3 µm superficially porous particle packed column with charged aerosol detection for the simple and sensitive measurement of nine macrolides in human urine.

Due to the lack of chromophores in many macrolides, analytical methods based on mass spectrometry and electrochemical detection coupled to liquid chromatography have been suggested to be suitable for the quantification of macrolides in complex matrices. In this study, a simple and sensitive analytical method was established for the simultaneous measurement of nine macrolides in human urine by combining a sub-3 µm superficially porous particle packed column with charged aerosol detection. After thorough investigation of various sample preparation methods, including two liquid-liquid extraction methods and four solid-phase extraction methods, HLB solid-phase extraction was selected and further optimized. Absolute recovery of the optimized sample preparation method ranged from 99.5-110.2%, indicating its very high extraction/clean-up efficiency. For chromatography, parameters influencing macrolide separation were systematically optimized, and the resulting conditions allowed baseline separation of nine macrolides within 24 min using a very simple mobile phase. The established method was validated for linearity, limit of detection, limit of quantification, absolute recovery, and precision. Based on its limit of detection (0.025-0.100 µg/mL), the method had similar or greater sensitivity than most methods based on electrochemical detection. It was found that the current method was appropriate for application to real human urine samples after drug administration.

Evaluation of four different analytical tools to determine the regional origin of Gastrodia elata and Rehmannia glutinosa on the basis of metabolomics study.

Chemical profiles of medicinal plants could be dissimilar depending on the cultivation environments, which may influence their therapeutic efficacy. Accordingly, the regional origin of the medicinal plants should be authenticated for correct evaluation of their medicinal and market values. Metabolomics has been found very useful for discriminating the origin of many plants. Choosing the adequate analytical tool can be an essential procedure because different chemical profiles with different detection ranges will be produced according to the choice. In this study, four analytical tools, Fourier transform near­infrared spectroscopy (FT-NIR), 1H-nuclear magnetic resonance spectroscopy (1H­NMR), liquid chromatography-mass spectrometry (LC-MS), and gas chromatography-mass spectroscopy (GC-MS) were applied in parallel to the same samples of two popular medicinal plants (Gastrodia elata and Rehmannia glutinosa) cultivated either in Korea or China. The classification abilities of four discriminant models for each plant were evaluated based on the misclassification rate and Q2 obtained from principal component analysis (PCA) and orthogonal projection to latent structures-discriminant analysis (OPLS­DA), respectively. 1H-NMR and LC-MS, which were the best techniques for G. elata and R. glutinosa, respectively, were generally preferable for origin discrimination over the others. Reasoned by integrating all the results, 1H-NMR is the most prominent technique for discriminating the origins of two plants. Nonetheless, this study suggests that preliminary screening is essential to determine the most suitable analytical tool and statistical method, which will ensure the dependability of metabolomics-based discrimination.

Ginsenosides Decrease ß-Amyloid Production via Potentiating Capacitative Calcium Entry.

Alzheimer's disease (AD) is a progressive and irreversible neurodegenerative disorder characterized by extracellular amyloid plaques composed of amyloid ß-peptide (Aß). Studies have indicated that Ca2+ dysregulation is involved in AD pathology. It is reported that decreased capacitative Ca2+ entry (CCE), a refilling mechanism of intracellular Ca2+, resulting in increased Aß production. In contrast, constitutive activation of CCE could decrease Aß production. Panax ginseng Meyer is known to enhance memory and cognitive functions in healthy human subjects. We have previously reported that some ginsenosides decrease Aß levels in cultured primary neurons and AD mouse model brains. However, mechanisms involved in the Aß-lowering effect of ginsenosides remain unclear. In this study, we investigated the relationship between CCE and Aß production by examining the effects of various ginsenosides on CCE levels. Aß-lowering ginsenosides such as Rk1, Rg5, and Rg3 potentiated CCE. In contrast, ginsenosides without Aß-lowering effects (Re and Rb2) failed to potentiate CCE. The potentiating effect of ginsenosides on CCE was inhibited by the presence of 2-aminoethoxydipherryl borate (2APB), an inhibitor of CCE. 2APB alone increased Aß42 production. Furthermore, the presence of 2APB prevented the effects of ginsenosides on Aß42 production. Our results indicate that ginsenosides decrease Aß production via potentiating CCE levels, confirming a close relationship between CCE levels and Aß production. Since CCE levels are closely related to Aß production, modulating CCE could be a novel target for AD therapeutics.

Sesquiterpenoids and hexanorcucurbitacin from Aquilaria malaccensis agarwood with anti-inflammatory effects by inhibiting the STAT1/AKT/MAPK/NLRP3 pathway.

Seven unknown compounds 1-7, including four sesquiterpenoids, one azulene-type, one indene-type, and one rare hexanorcucurbitacin, together with eleven knowns ones (8-16), were isolated from the agarwood chips of Aquilaria malaccensis. The structures of the isolated compounds were elucidated by extensive spectroscopic methods such as mass spectrometry, UV, IR, NMR spectroscopy. The precise stereo-chemical configurations of new compounds were determined by calculated ECD spectra data, as well as a single-crystal X-ray diffraction analysis. The isolated compounds 1-7 were evaluated by estimating the levels of nitric oxide (NO), TNF-α, and the expression of enzyme iNOS, and COX-2. Among them, a rare hexanortriterpenoid (7) derived from a cucurbitane-type triterpenoid showed the significantly attenuated neuro-inflammatory effects via the STAT1/AKT/MAPK/NLRP3 signaling pathway on the mechanistic studies.

Characterization of the structure, anti-inflammatory activity and molecular docking of a neutral polysaccharide separated from American ginseng berries.

AIM: American ginseng berries, grown in the aerial parts and harvested in August, are a potentially valuable material. The aim of the study was to analyze the specific polysaccharides in American ginseng berries, and to demonstrate the anti-inflammation effect through in vitro and in vivo experiments and molecular docking. METHODS: After deproteinization and dialysis, the extracted crude polysaccharide was separated and purified. The structure of the specific isolated polysaccharide was investigated by Fourier Transform infrared spectroscopy (FT-IR), GC-MS and nuclear magnetic resonance (NMR), and anti-inflammatory activity was evaluated using in vitro and in vivo models (Raw 264.7 cells and zebrafish). Molecular docking was used to analyze the binding capacity and interaction with cyclooxygenase-2 (COX-2). RESULTS: A novel neutral polysaccharide fraction (AGBP-A) was isolated from American ginseng berries. The structural analysis demonstrated that AGBP-A had a weight-average molecular weight (Mw) of 122,988 Da with a dispersity index (Mw/Mn) value of 1.59 and was composed of arabinose and galactose with a core structure containing →6)-Gal-(1→ residues as the backbone and a branching substitution at the C3 position. The side-chains comprised of α-L-Ara-(1→, α-L-Ara-(1→, →5)-α-L-Ara-(1→, ß-D-Gal-(1→. The results showed that it significantly decreased pro-inflammatory cytokines in the cell model. In a zebrafish model, AGBP-A reduced the massive recruitment of neutrophils to the caudal lateral line neuromast, suggesting the relief of inflammation. Molecular docking was used to analyze the combined capacity and interaction with COX-2. CONCLUSION: Our study indicated the potential efficacy of AGBP-A as a safe and valid natural anti-inflammatory component.

Discrimination between genetically identical peony roots from different regions of origin based on 1H-nuclear magnetic resonance spectroscopy-based metabolomics: determination of the geographical origins and estimation of the mixing proportions of blended samples.

Sixty peony root training samples of the same age were collected from various regions in Korea and China, and their genetic diversity was investigated for 23 chloroplast intergenic space regions. All samples were genetically indistinguishable, indicating that the DNA-based techniques employed were not appropriate for determining the samples' regions of origin. In contrast, (1)H-nuclear magnetic resonance ((1)H-NMR) spectroscopy-based metabolomics coupled with multivariate statistical analysis revealed a clear difference between the metabolic profiles of the Korean and Chinese samples. Orthogonal projections on the latent structure-discrimination analysis allowed the identification of potential metabolite markers, including γ-aminobutyric acid, arginine, alanine, paeoniflorin, and albiflorin, that could be useful for classifying the samples' regions of origin. The validity of the discrimination model was tested using the response permutation test and blind prediction test for internal and external validations, respectively. Metabolomic data of 21 blended samples consisting of Korean and Chinese samples mixed at various proportions were also acquired by (1)H-NMR analysis. After data preprocessing which was designed to eliminate uncontrolled deviations in the spectral data between the testing and training sets, a new statistical procedure for estimating the mixing proportions of blended samples was established using the constrained least squares method for the first time. The predictive procedure exhibited relatively good predictability (adjusted R (2) = 0.7669), and thus has the potential to be used in the quality control of peony root by providing correct indications for a sample's geographical origins.