Discrimination of Lonicera japonica THUNB. from different geographical origins using restriction fragment length polymorphism analysis.

Lonicera japonica THUNB. is a commonly used anti-inflammatory herbal medicine. The therapeutic effectiveness of L. japonica depends significantly on its geographical origin. However, it is difficult to define criteria for confirming geographical authenticity using microscopic and chemical characteristics. In the present study, the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA loci of L. japonica from different origins and related species was sequenced. The mutation site found in the ITS region from geo-authentic L. japonica can be recognized by the restriction endonuclease EcoN I. Since PCR products from geo-authentic L. japonica cannot be digested completely, a quantitative restriction fragment length polymorphism analysis method was developed. The cleavage rate of PCR products by EcoN I was determined to be more than 70% in all geo-authentic L. japonica and less than 20% in non-geo-authentic L. japonica and other species from genus Lonicera. The rate correlated remarkably with the geographical origin of L. japonica. Therefore, this method can be used to classify geo-authentic L. japonica.

Identification of Fritillaria pallidiflora using diagnostic PCR and PCR-RFLP based on nuclear ribosomal DNA internal transcribed spacer sequences.

Fritillaria pallidiflora Schrenk (Liliaceae) is a commonly used antitussive herb. There are 9 species of Fritillaria recorded as herbal drugs in the Chinese Pharmacopoeia. The other species are often marketed as F. pallidiflora, and thus, the therapeutic effects of F. pallidiflora are not achieved. Methods to distinguish F. pallidiflora from the 8 other species of Fritillaria are limited by the current morphological and chemical methods. In this study, we report two molecular authentication methods based on the sequences of nuclear ribosomal DNA internal transcribed spacer (nrDNA ITS) regions. For diagnostic PCR, we designed a pair of species-specific primers to authenticate F. pallidiflora. The PCR program consisted of only two steps for every repeated cycle. For PCR-RFLP, we identified a distinctive site which can be recognized by the restriction endonuclease Eco81I in the nrDNA ITS1 region of F. pallidiflora. PCR-RFLP analysis was established to differentiate F. pallidiflora from the other species of Fritillaria. These methods provide effective and accurate identification of F. pallidiflora.

[Studies on influence of fungal elicitor on hairy root of Panax ginseng biosynthesis ginseng saponin and biomass].

OBJECTIVE: To study influence of fungal elicitors on the biomass and ginseng saponin biosynthesis of hairy roots of Panax ginseng (HRPG). METHOD: Fungal elicitors were extracted from Colletorichum lagnarinm, Phoma filtrate, Fusarium oxysponum, Asperillus niger and culture with HRPG. The total ginseng sponin and four kinds of monomeric sponins were analysed by UV-spectrophotometry and RP-HPLC. RESULT: Fungal elicitors coula not only can influence on HRPG biomass and total ginseng sponin, but also improve or decrease some monomeric sponin. The total ginseng sponin could be increased to 3.649% but Rg1 and Re could not be detected when A. niger elicitors wss 20 mg x L(-1) in the culture fluid. CONCLUSION: Fungal elicitor has specificity influence on secondary metabolite of HRPG. HRPG can biosynthesize specially active component by using specific fungal elicitor is used.