High-Performance Thin-Layer Chromatographic Fingerprints of Triterpenoids for Distinguishing Between Isodon lophanthoides and Isodon lophanthoides var. gerardianus.

Background: The aerial parts of Isodon lophanthoides (Buch. Ham. ex D. Don) Hara (IL) has been officially recorded as Isodonis lophanthoidis herba by many provincial quality control standards for Chinese herbal medicines in China. Recently, it has been found that one of its varieties, I. lophanthoides var. gerardianus (Benth.) Hara (ILVG) was pretended to be I. lophanthoidis herba in herbal material markets or cultivated bases. Because of the similarity on appearance, these two close-related species were difficult to be identified by morphological characters, especially when they are dried and sliced. Objective: To establish a rapid and specific method for identification of the two herbal medicines. Method: In this paper, a fingerprint of triterpenoids by HPTLC coupled with a digital profiling was established to identify IL and distinguish it from its substitute, ILVG. The specific HPTLC fingerprints constructed by determining twelve batches of IL samples and thirteen batches of ILVG samples, intuitionally reflected the difference between the two species on HPTLC image and the peak-peak rations of chemical distribution. Results: Authentication results of nine batches of commercial samples by the above established HPTLC fingerprints exhibited coincident conclusion with that by morphological means. Conclusions: The HPTLC fingerprint is proven to be simple, repeatable, specific, and suitable for rapid identification of I. lophanthoidis herba. Highlights: An efficient method for identification and distinguishing Isodon lophanthoides from its substitute, I. lophanthoides var. gerardianus, was established. HPTLC fingerprints of ursane-type triterpenoides were constructed and validated by determining IL and ILVG samples.

Phylogeny and historical biogeography of Isodon (Lamiaceae): rapid radiation in south-west China and Miocene overland dispersal into Africa.

Rapid organismal radiations occurring on the Qinghai-Tibetan Plateau (QTP) and the mechanisms underlying Asia-Africa intercontinental disjunctions have both attracted much attention from evolutionary biologists. Here we use the genus Isodon (Lamiaceae), a primarily East Asian lineage with disjunct species in central and southern Africa, as a case study to shed light upon these processes. The molecular phylogeny and biogeographic history of Isodon were reconstructed using sequences of three plastid markers, the nuclear ribosomal internal transcribed spacer (nrITS), and a low-copy nuclear gene (LEAFY intron II). The evolution of chromosome numbers in this genus was also investigated using probabilistic models. Our results support a monophyletic Isodon that includes the two disjunct African species, both of which likely formed through allopolyploidy. An overland migration from Asia to Africa through Arabia during the early Miocene is proposed as the most likely explanation for the present disjunct distribution of Isodon. The opening of the Red Sea in the middle Miocene may appear to have had a major role in disrupting floristic exchange between Asia and Africa. In addition, a rapid radiation of Isodon was suggested to occur in the late Miocene. It corresponds with one of the major uplifts of the QTP and subsequent aridification events. Our results support the hypothesis that geological and climatic events play important roles in driving biological diversification of organisms distributed in the QTP area.

Investigations of anticholinestrase and antioxidant potentials of methanolic extract, subsequent fractions, crude saponins and flavonoids isolated from Isodon rugosus.

BACKGROUND: Based on the ethnomedicinal uses and the effective outcomes of natural products in various diseases, this study was designed to evaluate Isodon rugosus as possible remedy in oxidative stress, alzheimer's and other neurodegenerative diseases. Acetylecholinestrase (AChE) and butyrylcholinesterase (BChE) inhibitory activities of crude methanolic extract (Ir.Cr), resultant fractions (n-hexane (Ir.Hex), chloroform (Ir.Cf), ethyl acetate (Ir.EtAc), aqueous (Ir.Aq)), flavonoids (Ir.Flv) and crude saponins (Ir.Sp) of I. rugosus were investigated using Ellman's spectrophotometric method. Antioxidant potential of I. rugosus was determined using DPPH, H2O2 and ABTS free radicals scavenging assays. Total phenolic and flavonoids contents of plant extracts were determined and expressed in mg GAE/g dry weight and mg RTE/g of dry sample respectively. RESULTS: Among different fractions Ir.Flv and Ir.Cf exhibited highest inhibitory activity against AChE (87.44 ± 0.51, 83.73 ± 0.64%) and BChE (82.53 ± 0.71, 88.55 ± 0.77%) enzymes at 1 mg/ml with IC50 values of 45, 50 for AChE and 40, 70 µg/ml for BChE respectively. Activity of these fractions were comparable to galanthamine causing 96.00 ± 0.30 and 88.61 ± 0.43% inhibition of AChE and BChE at 1 mg/ml concentration with IC50 values of 20 and 47 µg/ml respectively. In antioxidant assays, Ir.Flv, Ir.Cf, and Ir.EtAc demonstrated highest radicals scavenging activities in DPPH and H2O2 assays which were comparable to ascorbic acid. Ir.Flv was found most potent with IC50 of 19 and 24 µg/ml against DPPH and H2O2 radicals respectively. Whereas antioxidant activates of plant samples against ABTS free radicals was moderate. Ir.Cf, Ir.EtAc and Ir.Cr showed high phenolic and flavonoid contents and concentrations of these compounds in different fractions correlated well to their antioxidant and anticholinestrase activities. CONCLUSION: It may be inferred from the current investigations that the Ir.Sp, Ir.Flv and various fractions of I. rugosus are good sources of anticholinesterase and antioxidant compounds. Different fractions can be subjected to activity guided isolation of bioactive compounds effective in neurological disorders.

Investigations of anticholinestrase and antioxidant potentials of methanolic extract, subsequent fractions, crude saponins and flavonoids isolated from Isodon rugosus

BACKGROUND: Based on the ethnomedicinal uses and the effective outcomes of natural products in various diseases, this study was designed to evaluate Isodon rugosus as possible remedy in oxidative stress, alzheimer's and other neurodegenerative diseases. Acetylecholinestrase (AChE) and butyrylcholinesterase (BChE) inhibitory activities of crude methanolic extract (Ir.Cr), resultant fractions (n-hexane (Ir.Hex), chloroform (Ir.Cf), ethyl acetate (Ir.EtAc), aqueous (Ir.Aq)), flavonoids (Ir.Flv) and crude saponins (Ir.Sp) of I. rugosus were investigated using Ellman's spectrophotometric method. Antioxidant potential of I. rugosus was determined using DPPH, H2O2 and ABTS free radicals scavenging assays. Total phenolic and flavonoids contents of plant extracts were determined and expressed in mg GAE/g dry weight and mg RTE/g of dry sample respectively. RESULTS: Among different fractions Ir.Flv and Ir.Cf exhibited highest inhibitory activity against AChE (87.44 ± 0.51, 83.73 ± 0.64%) and BChE (82.53 ± 0.71, 88.55 ± 0.77%) enzymes at 1 mg/ml with IC50 values of 45, 50 for AChE and 40, 70 µg/ml for BChE respectively. Activity of these fractions were comparable to galanthamine causing 96.00 ± 0.30 and 88.61 ± 0.43% inhibition of AChE and BChE at 1 mg/ml concentration with IC50 values of 20 and 47 µg/ml respectively. In antioxidant assays, Ir.Flv, Ir.Cf, and Ir.EtAc demonstrated highest radicals scavenging activities in DPPH and H2O2 assays which were comparable to ascorbic acid. Ir.Flv was found most potent with IC50 of 19 and 24 µg/ml against DPPH and H2O2 radicals respectively. Whereas antioxidant activates of plant samples against ABTS free radicals was moderate. Ir.Cf, Ir.EtAc and Ir.Cr showed high phenolic and flavonoid contents and concentrations of these compounds in different fractions correlated well to their antioxidant and anticholinestrase activities. CONCLUSION: It may be inferred from the current investigations that the Ir.Sp, Ir.Flv and various fractions of I. rugosus are good sources of anticholinesterase and antioxidant compounds. Different fractions can be subjected to activity guided isolation of bioactive compounds effective in neurological disorders.

Selection for high oridonin yield in the Chinese medicinal plant Isodon (Lamiaceae) using a combined phylogenetics and population genetics approach.

Oridonin is a diterpenoid with anti-cancer activity that occurs in the Chinese medicinal plant Isodon rubescens and some related species. While the bioactivity of oridonin has been well studied, the extent of natural variation in the production of this compound is poorly known. This study characterizes natural variation in oridonin production in order to guide selection of populations of Isodon with highest oridonin yield. Different populations of I. rubescens and related species were collected in China, and their offspring were grown in a greenhouse. Samples were examined for oridonin content, genotyped using 11 microsatellites, and representatives were sequenced for three phylogenetic markers (ITS, rps16, trnL-trnF). Oridonin production was mapped on a molecular phylogeny of the genus Isodon using samples from each population as well as previously published Genbank sequences. Oridonin has been reported in 12 out of 74 species of Isodon examined for diterpenoids, and the phylogeny indicates that oridonin production has arisen at least three times in the genus. Oridonin production was surprisingly consistent between wild-collected parents and greenhouse-grown offspring, despite evidence of gene flow between oridonin-producing and non-producing populations of Isodon. Additionally, microsatellite genetic distance between individuals was significantly correlated with chemical distance in both parents and offspring. Neither heritability nor correlation with genetic distance were significant when the comparison was restricted to only populations of I. rubescens, but this result should be corroborated using additional samples. Based on these results, future screening of Isodon populations for oridonin yield should initially prioritize a broad survey of all species known to produce oridonin, rather than focusing on multiple populations of one species, such as I. rubescens. Of the samples examined here, I. rubescens or I. japonicus from Henan province would provide the best source of oridonin.

[Identification of herba rabdosiae serrae from different plant resources by RAPD method].

OBJECTIVE: To identify the different plant resources of Herba Rabdosiae Serrae by using Random Amplified Polymorphic DNA ( RAPD ) Analysis. METHODS: The mini spin columns were used to extract the genomic DNA from five different plants of Herba Rabdosiae Serrae. With the DNA extracted from these plants as template,the 85 oligo nucleic acids (10 bp)as random primers,the polymer chain reaction (PCR) was done and the results were analysed by electro-pharoses. RESULTS: 12 primers were selected with polymorphism and 7 of them showed good polymorphism in RAPD map. CONCLUSION: RAPD method can be used to identify the plant resources of Herba Rabdosiae Serrae.

[IR-fingerprinting ordination comparison on different organs of three species of isodon].

The plants of the genus Isodon are of important medicinal values, being widely used in the production of the Chinese traditional and herbal drugs. Compared with the other identification methods, the identification of the Chinese traditional and herbal drugs using the fourier-transform infrared spectrometer with OMNI collector is simple and convenient, fast and accurate. Moreover, the extraction or break of the samples are not necessary in the identification using OMNI collector. In the present paper, fourier-transform infrared spectrometer with OMNI collector is applied to gain the IR-fingerprintings of eleven samples of Isodon inflexus, I. Lophanthoides and I. Macrocalyx. Based on the indices of wavenumber-absorbance, the differences of eleven IR-fingerprintings are compared by PCA (Principal Component Analysis). The results show that it is practical to apply PCA on the basis of IR-fingerprinting to compare the chemical differences of plant samples.

[A study on quality standard of Rabdosia lophanthoides (Buch.-Ham. ex D. Don Hara)].

OBJECTIVE: To establish quality standards for safe and correct use of Rabdosia lophanthoides. METHOD: Field investigation and pharmacognostic study. RESULT: R. lophanthoides was confirmed as a derivative of R. lophanthoides. CONCLUSION: A convenient and effective method for identifying the herb has been established.