HPLC-DAD Fingerprints Combined With Multivariate Analysis of Epimedii Folium From Major Producing Areas in Eastern Asia: Effect of Geographical Origin and Species

Abstract

The growth location and plant variety may influence the active components and biological activities of plants used in phytomedicine. In this study, nine sets of different Epimedii Folium, from different representative cultivation locations and Epimedium species, were collected for comparison, using HPLC-DAD combined with multivariate analysis. The objective was to investigate the influence of geographical origin and Epimedium species on the quality of Epimedii Folium, and provide applicable guidance for cultivation and quality control of Epimedii Folium. Several Epimedium spp. sets were used to establish the HPLC-DAD fingerprints and 91 peaks (compounds) were selected for the multivariate analysis. Major compounds were analyzed by HPLC-DAD combined with principal component analysis (PCA). HPLC quantitative analysis of known bioactive compounds was performed. Application of PCA to HPLC data showed that Epimedium samples sharing the same geographical origin or species clustered together, indicating that both species and geographical origin have impacts on the quality of Epimedii Folium. The major bioactive flavonoid compounds, epimedin C, icariin and baohuoside I, were identified and quantified. The concentration of bioactive compounds was significantly influenced both by species and geographical origin. E. sagittatum from Sichuan showed the highest content of bioactive compounds. The results showed that both Epimedium species and geographical origin have strong impact into quality of Epimedii Folium. HPLC data combined with multivariate analysis is a suitable approach to inform the selection of cultivation areas and choose Epimedium spp. most suitable for different geographical areas, resulting in improved quality of Epimedii Folium.

Keywords: Epimedium sp; baohuoside I; epimedin C; high performance liquid chromatography (HPLC); icariin; principal component analysis (PCA).

FIGURE 1

Geographical distribution of Epimedium samples collected in this study.

FIGURE 2

HPLC fingerprints of nine sets of Epimedium samples studied and standards (epimedin C, icariin and baohuoside I, from left to right).

FIGURE 3

PCA scores plot (A) and loadings plot (B) of Epimedium samples. In the scores plot, Epimedium species are represented by different shapes, and each data point is colored according to its amount of epimedin C and labeled with its geographical origin (SK—South Korea, LG—Longnan Gansu, WG—Weiyuan Gansu, Ji—Jilin, Si—Sichuan, Sh—Shaanxi). Ellipses indicate sample groups based on species or geographical origin; dashed-line circles indicate outliers (two E. brevicornum outliers from Weiyuan Gansu, one E. sagittatum outlier and another E. pubescens both from Sichuan). The loadings plot (B) shows the compounds responsible for the group separation; for clarity, the peak numbers corresponding to epimedin C, icariin and baohuoside I were substituted by their name.

FIGURE 4

PCA scores plot (A) for same Epimedium species (E. koreanum, EK) collected in different locations (Daqiu, South Korea and Jilin, China)—A; and PCA scores plot (B) for different species (E. sagittatum, EK, and E. pubescens, EP) cultivated in the same field (Wanyuan, Sichuan).

FIGURE 5

Amount (mg/g dry weight biomass) of epimedin C, icariin and baohuoside I in E. pubescens (A), E. brevicornum (B) and E. koreanum (C) cultivated in different geographical locations. Bars represent average ±SEM; dots represent individual data points. Statistically significant differences are represented by asterisks (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001).

FIGURE 6

Amount (mg/g dry weight biomass) of epimedin C, icariin and baohuoside I of different plant species cultivated in same place (Wanyuan, Sichuan). Bars represent average ±SEM; dots represent individual data points. Statistically significant differences are represented by asterisks (**p < 0.01, ****p < 0.0001).