| Drinking herbal tea is more and more popular in China and Southeast Asian Nations. However, study on the species identification and quality evaluation of the commonly used herbal tea materials is insufficient.;In this study, the first step was to collect multiple samples of Herba Oldenlandiae and its substitutes from different growing areas. The second step was to establish the characteristics by which the three species of Oldenlandia under investigation could be distinguished.;Sequences of internal transcribed spacer (ITS) were also used to identify Herba Oldenlandiae and its substitutes. According to the alignment of those sequences, thirteen position-specific nucleotides were found in the ITS sequences of O. diffusa which could be used for identification Herba Oldenlandiae and its substitutes. In addition, the phylogenetic tree based on the tested ITS sequences was reconstructed. The results showed that O. diffusa had a close relationship to O. tenelliflora but not to O. corymbosa.;The antiproliferative effect of O. diffusa and its substitutes, O. corymbosa and O. tenelliflora, on human colon carcinoma CaCo2 cells and hepatoma HepG2 cells was analyzed. The SRB assay on human hepatoma HepG2 cells and human colon carcinoma CaCo2 cells was used for this purpose. The results showed that there was almost no antiproliferative effect of the methanol extracts from O. diffusa, O. corymbosa and O. tenelliflora, while the chloroform extracts from O. diffusa and O. corymbosa exhibited slightly antiproliferative effects. Further investigation on the antiproliferative constituents of O. diffusa led to the isolation of a new compound, E-6- O-p-coumaroyl scandoside methyl ester-10-O-methyl ether (54), together with six known compounds---asperuloside (1), E-6-O-p-coumaroyl scandoside methyl ester (3), oleanolic acid (22), ursolic acid (23), beta-sitosterol glucoside (51) and betulin (53). Among the above compounds, compound 53 was firstly isolated from this herb, and compound 54 was a new iridoid glucoside. The results also indicated that the chloroform fraction of O. diffusa contained a great quantity of ursolic acid. Therefore, ursolic acid was considered to be the main antiproliferative constituent.;The differences in the constituents of Herba Oldenlandiae and its substitutes were analyzed by HPLC fingerprints and LC-HR-ESI-MS methods.;As oleanolic acid and ursolic acid are the anti-tumor components of O. diffusa reported in the literatures and as there is no systematic study comparing the contents of these constituents in multiple samples of the three species used as Herba Oldenlandiae, the parameters were attempted to establish by which these two components could be used to evaluate the quality of Herba Oldenlandiae and its substitutes. A convenient method with good resolution was developed. The results showed that the contents of oleanolic acid and ursolic acid in O. diffusa were generally lower by almost two times than in O. corymbosa. Therefore, quantifying these constituents could readily and reliably distinguish O. diffusa from O. corymbosa---but not from O. tenelliflora because the contents of these two compounds in this species were similar to those of O. diffusa.;In conclusion, the techniques of fluorescence microscopy and DNA molecular markers were used to distinguish Herba Oldenlandiae and its substitutes. The chemicals of O. diffusa, O. corymbosa and O. tenelliflora were quite different based on the above analysis. All the results indicated that the three herbs should not be used interchangeably. Moreover, the level of iridoid glucosides in the dry and fresh herbal samples was significantly different. These studies successfully addressed the problem of species identification and quality evaluation of Herba Oldenlandiae. (Abstract shortened by UMI.)... |
