Chemical Relationship among Genetically Authenticated Medicinal Species of Genus Angelica.

The genus Angelica comprises various species utilized for diverse medicinal purposes, with differences attributed to the varying levels or types of inherent chemical components in each species. This study employed DNA barcode analysis and HPLC analysis to genetically authenticate and chemically classify eight medicinal Angelica species (n = 106) as well as two non-medicinal species (n = 14) that have been misused. Nucleotide sequence analysis of the nuclear internal transcribed spacer (ITS) region revealed differences ranging from 11 to 117 bp, while psbA-trnH showed variances of 3 to 95 bp, respectively. Phylogenetic analysis grouped all samples except Angelica sinensis into the same cluster, with some counterfeits forming separate clusters. Verification using the NCBI database confirmed the feasibility of species identification. For chemical identification, a robust quantitative HPLC analysis method was developed for 46 marker compounds. Subsequently, two A. reflexa-specific and seven A. biserrata-specific marker compounds were identified, alongside non-specific markers. Moreover, chemometric clustering analysis reflecting differences in chemical content between species revealed that most samples formed distinct clusters according to the plant species. However, some samples formed mixed clusters containing different species. These findings offer crucial insights for the standardization and quality control of medicinal Angelica species.

Chemotaxonomic Classification of Peucedanum japonicum and Its Chemical Correlation with Peucedanum praeruptorum, Angelica decursiva, and Saposhnikovia divaricata by Liquid Chromatography Combined with Chemometrics.

The roots of Peucedanum japonicum (Apiaceae) have been used as an alternative to the roots of Saposhnikovia divaricata (Apiaceae) to treat common cold-related symptoms in Korea. However, a variety of Peucedanum species, including the roots of P. praeruptorum or Angelica decursiva (=P. decursivum), have been used to treat phlegm-heat-induced symptoms in China. Hence, as there are differences in the medicinal application of P. japonicum roots between Korea and China, chemotaxonomic classification of P. japonicum was evaluated. Sixty samples derived from P. japonicum, P. praeruptorum, A. decursiva, and S. divaricata were phylogenetically identified using DNA barcoding tools, and chemotaxonomic correlations among the samples were evaluated using chromatographic profiling with chemometric analyses. P. japonicum samples were phylogenetically grouped into the same cluster as P. praeruptorum samples, followed by S. divaricata samples at the next cluster level, whereas A. decursiva samples were widely separated from the other species. Moreover, P. japonicum samples showed higher chemical correlations with P. praeruptorum samples or A. decursiva samples, but lower or negative chemical correlations with S. divaricata samples. These results demonstrate that P. japonicum is more genetically and chemically relevant to P. praeruptorum or A. decursiva and, accordingly, the medicinal application of P. japonicum might be closer to the therapeutic category of these two species than that of S. divaricata.

Neuroprotective effects of ethanolic extract from dry Rhodiola rosea L. rhizomes.

Rhodiola rosea L. rhizome has been used as a traditional medicine to treat fatigue, depression, and cognitive dysfunction. We aimed to authenticate R. rosea L. rhizome using the DNA barcoding technique and to quantify its main compounds, total phenolics, total flavonoids, and antioxidant capacity, and then to investigate their neuroprotective effects. The sequences of internal transcribed spacer and trnH-psbA of R. rosea L. rhizomes showed a 99% identity with those of NCBI GenBank database according to BLAST searches. Analysis using reversed-phase HPLC revealed five main compounds in R. rosea L. rhizome. Rhodiola rosea L. rhizome and two bioactive compounds, salidroside and tyrosol, showed free radical scavenging activity. Rhodiola rosea L. rhizome and its identified compounds protected neuronal PC-12 cells against oxidative stress and showed moderate acetylcholinesterase inhibition. Taken together, these results suggest that R. rosea L. rhizomes with bioactives can be used as a functional ingredient with potential for neuroprotection. SUPPLEMENTARY INFORMATION: The online version of this article (doi:10.1007/s10068-020-00868-7) contains supplementary material, which is available to authorized users.

Aqueous extract of Chrysanthemum morifolium Ramat. inhibits RANKL-induced osteoclast differentiation by suppressing the c-fos/NFATc1 pathway.

OBJECTIVE: The flower of chrysanthemum, used worldwide as a medicinal and edible product, has shown various bioactivities, such as anti-inflammatory, antioxidant, anti-tumorigenic, and hepatoprotective activities, as well as cardiovascular protection. However, the effect of Chrysanthemum morifolium Ramat. on the regulation of osteoclast differentiation has not yet been reported. In this study, we aimed to investigate the inhibitory effect of Chrysanthemum morifolium Ramat. water extract (CME) on RANKL-induced osteoclast differentiation in mouse bone marrow-derived macrophages (BMMs). STUDY DESIGN: Bone marrow-derived macrophages (BMMs) isolated from the C57BL/6 J mice. The viability of BMMs was detected with MTT assays. Inhibitory effects of CME on osteoclast differentiation and bone resorption was measured by TRAP staining and Pit assay. Osteoclast differentiation-associated gene expression were assessed by Real-time quantitative polymerase chain reaction. Intracellular signaling molecules was assessed by western blot. RESULTS: CME significantly inhibited osteoclast differentiation in BMMs without cytotoxicity, besides inhibiting MAPK/c-fos and PLCγ2/CREB activation. The inhibitory effects of CME on differentiation-related signaling molecules resulted in significant repression of NFATc1 expression, which is a key transcription factor in osteoclast differentiation, fusion, and activation. CONCLUSION: Our results confirmed the inhibition of RANKL-induced PLCγ2/CREB/c-fos/NFATc1 activation by CME during osteoclast differentiation. The findings collectively suggested CME as a traditional therapeutic agent for osteoporosis, RA, and periodontitis.

Quantitative Comparison of the Marker Compounds in Different Medicinal Parts of Morus alba L. Using High-Performance Liquid Chromatography-Diode Array Detector with Chemometric Analysis.

It is thought that the therapeutic efficacy of Morus alba L. is determined by its biological compounds. We investigated the chemical differences in the medicinal parts of M. alba by analyzing a total of 57 samples (15 root barks, 11 twigs, 12 fruits, and 19 leaves). Twelve marker compounds, including seven flavonoids, two stilbenoids, two phenolic acids, and a coumarin, were quantitatively analyzed using a high-performance liquid chromatography-diode array detector and chemometric analyses (principal component and heatmap analysis). The results demonstrated that the levels and compositions of the marker compounds varied in each medicinal part. The leaves contained higher levels of six compounds, the root barks contained higher levels of four compounds, and the twigs contained higher levels of two compounds. The results of chemometric analysis showed clustering of the samples according to the medicinal part, with the marker compounds strongly associated with each part: mulberroside A, taxifolin, kuwanon G, and morusin for the root barks; 4-hydroxycinnamic acid and oxyresveratrol for the twigs and skimmin; chlorogenic acid, rutin, isoquercitrin, astragalin, and quercitrin for the leaves. Our approach plays a fundamental role in the quality evaluation and further understanding of biological actions of herbal medicines derived from various medicinal plant parts.

Chemotaxonomic Monitoring of Genetically Authenticated Amomi Fructus Using High-Performance Liquid Chromatography-Diode Array Detector with Chemometric Analysis.

Amomi Fructus is widely used to treat digestive disorders, and Amomum villosum, A. villosum var. xanthioides, and A. longiligulare are permitted medicinally in national pharmacopeias. However, there are a variety of adulterants present in herbal markets owing to their morphological similarities to the genuine Amomum species. Forty-two Amomi Fructus samples from various origins were identified using internal transcribed spacer and chloroplast barcoding analyses, and then their chromatographic profiles were compared using chemometric analysis for chemotaxonomic monitoring. Among the Amomi Fructus samples, A. villosum, A. longiligulare, A. ghaticum, and A. microcarpum were confirmed as single Amomum species, whereas a mixture of either these Amomum species or with another Amomum species was observed in 15 samples. Chemotaxonomic monitoring results demonstrated that two medicinal Amomum samples, A. villosum and A. longiligulare, were not clearly distinguished from each other, but were apparently separated from other non-medicinal Amomum adulterants. A. ghaticum and A. microcarpum samples were also chemically different from other samples and formed their own species groups. Amomum species mixtures showed diverse variations of chemical correlations according to constituent Amomum species. Genetic authentication-based chemotaxonomic monitoring methods are helpful in classifying Amomi Fructus samples by their original species and to distinguish genuine Amomum species from the adulterants.

DNA Markers to Discriminate Cannabis sativa L. 'Cheungsam' with Low Tetrahydrocannabinol (THC) Content from Other South Korea Cultivars Based on the Nucleotide Sequences of Tetrahydrocannabinolic Acid Synthase and Putative 3-Ketoacyl-CoA Synthase Genes.

Cannabis sativa L. has been utilized for a long time as a traditional herbal medicine in Korea. Dry fruits, achenes, each containing a single seed of Cannabis, are currently prescribed as Ma In (Cannabis Semen), a laxative. As each achene is enclosed by a bract, in which tetrahydrocannabinol (THC), the main psychological active compound in Cannabis is synthesized; achene is easily contaminated by THC from bract remnants. Therefore, it is safer to harvest achenes from Cannabis with a low THC content. Seeds of hemp, a low THC Cannabis, were recently classified as possible sources of new pharmacologically active compounds. Thus, a proper method to select appropriate Cannabis plants with low THC among cultivars in South Korea for medicinal purpose is necessary. As a result of cross-selection, Cannabis L. cultivar "Cheungsam" (CH) with the lowest THC content among cultivars cultivated in South Korea has been developed. In this study, we developed two DNA markers to reliably discriminate CH from other local cultivars with higher THC contents. We developed primer sets CHF3/CHR2 to amplify the 642 bp DNA marker of CH based on differences in the nucleotide sequences of the THCA synthase gene, which encodes a key enzyme in THC synthesis. We then developed a CHF1/CHR3 primer set to amplify the 401 bp DNA marker of CH based on the differences in both the content of very long chain fatty acids (VLCFs) and the sequence of the putative 3-ketoacyl-CoA synthase (KCS) gene encoding enzymes synthesizing VLCFs among local cultivars.

Identification and Monitoring of Amomi Fructus and its Adulterants Based on DNA Barcoding Analysis and Designed DNA Markers.

Amomi Fructus is one of the traditional medicines derived from the ripe fruits of the Zingiberaceae family of plants, which include Amomum villosum, A. villosum var. xanthioides, and A. longiligulare. Owing to their highly similar morphological traits, several kinds of adulterants of Amomi Fructus have been reported. Therefore, accurate and reliable methods of identification are necessary in order to ensure drug safety and quality. We performed DNA barcoding using five regions (ITS, matK, rbcL, rpoB, and trnL-F intergenic spacer) of 23 Amomi Fructus samples and 22 adulterants. We designed specific DNA markers for Amomi Fructus based on the single nucleotide polymorphisms (SNPs) in the ITS. Amomi Fructus was well separated from the adulterants and was classified with the species of origin based on the detected SNPs from the DNA barcoding results. The AVF1/ISR DNA marker for A. villosum produced a 270 bases amplified product, while the ALF1/ISF DNA marker produced a 350 bases product specific for A. longiligulare. Using these DNA markers, the monitoring of commercially distributed Amomi Fructus was performed, and the monitoring results were confirmed by ITS analysis. This method identified samples that were from incorrect origins, and a new species of adulterant was also identified. These results confirmed the accuracy and efficiency of the designed DNA markers; this method may be used as an efficient tool for the identification and verification of Amomi Fructus.

Quantitative Interrelation between Atractylenolide I, II, and III in Atractylodes japonica Koidzumi Rhizomes, and Evaluation of Their Oxidative Transformation Using a Biomimetic Kinetic Model.

Analytical methods based on ultraperformance liquid chromatography/ion-trap mass spectrometry (UPLC/ion-trap MS) were developed for quantification of atractylenolide I, II, and III in the methanol extract of Atractylodes japonica rhizomes with a C18 column in an acidified water/acetonitrile gradient eluent in an LC system, and ion-trap MS coupled with electrospray ionization was employed under positive-ion mode. The three atractylenolides were quantified in all A. japonica samples, and the content of atractylenolide I, II, and III showed a significant correlation to each other. Such high correlation was explained by the mechanistic insights into the biosynthetic pathway of atractylenoide III and I from atractylenoide II by using the biomimetic cytochrome P450 model, [Fe(tmp)](CF3SO3) (tmp = meso-tetramesitylporphyrin). Atractylenolides could be transformed by oxidation via the oxidative enzyme in the A. japonica plant. The present study first reports the first oxidative transformation of atractylenolides using the heme iron model complex.

Chemical Differentiation of Genetically Identified Atractylodes japonica, A. macrocephala, and A. chinensis Rhizomes Using High-Performance Liquid Chromatography with Chemometric Analysis.

The rhizome of Atractylodes japonica, which is a herbal medicine used for gastrointestinal therapeutics, has been categorized with A. macrocephala rhizome or A. chinensis rhizome based on different therapeutic criteria in Korea, China, and Japan. In the present study, 61 A. japonica, A. macrocephala, and A. chinensis rhizomes were collected from Korea and China and were genetically identified by internal transcribed spacer sequencing analysis. Chromatographic profiles were obtained from high-performance liquid chromatography analysis of the methanol and hot-water extracts of Atractylodes rhizomes and chemical differentiation of the rhizomes was carried out using chemometric statistical analyses such as principal component analysis, hierarchical clustering analysis, and Pearson's correlation coefficient analysis. The results from chromatographic profiles and chemometric analyses demonstrate that A. japonica rhizomes showed apparent chemical differences from A. macrocephala and A. chinensis rhizomes in the methanol extracts. In contrast, no clear distinction was apparent for the hot-water extracts of Atractylodes rhizomes, especially A. chinensis rhizomes. These results indicate that there is a clear chemical difference between A. japonica and A. macrocephala rhizomes; however, the chemical diversity of A. chinensis rhizome shows different chemical relationships with A. japonica or A. macrocephala rhizome, dependent on the chemical features.

Optimal Processing Conditions of Boswellia carteri Birdw. Using Response Surface Methodology.

BACKGROUND: Boswellia carteri Bridw. is being widely used for its anti-inflammatory properties, as well as for wound healing, antimicrobial, and immunomodulatory properties, and boswellic acids (BAs) are considered to be the main active constituents. OBJECTIVES: To investigate optimal conditions of stir-baking process for the resin of B. carteri with vinegar of using response surface methodology (RSM). MATERIALS AND METHODS: The concentration of acetic acid, heating temperature, and heating time were set as influential factors, and the yields of chemical compounds were the response values which were optimally designed by a Box-Behnken design. The amounts of 11-keto-ß-boswellic acid (KBA) and α-boswellic acid (αBA) in B. carteri resin were quantified using high-performance liquid chromatography analysis. RESULTS: Maximum amounts of KBA and αBA in B. carteri resin were obtained using 6% acetic acid for 10 min at 90°C in preliminary test. Two factor interactions, such as acetic acid concentration-heating temperature and heating temperature-heating time, were significantly observed by multiple regression analysis. Optimal processing conditions from RSM were 5.83% for acetic acid concentration, 9.56 min for heating time, and 89.87°C for heating temperature. Under the modified conditions, the experimental value of the response was 11.25 mg/g, which was similar to the predicted value. CONCLUSIONS: The results suggest that the optimal conditions for the stir-baking process of B. carteri resin were determined by RSM, which was reliable and applicable to practical processing of herbal medicine. SUMMARY: The resin of Boswellia carteri was macerated in aqueous acetic acid and heated using an oven for stir baking processThe interaction between heating temperature and heating time was the most significantOptimal conditions for processing B. carteri resin were determined as 5.83% acetic acid, 9.56 min for heating time, and 89.87°C for heating temperature. Abbreviations used: BAs: Boswellic acids; KBA: 11 keto ß boswellic acid; αBA: α boswellic acid; BBD: Box-Behnken design; RSM: Response surface method; HPLC: High performance liquid chromatography; LOD: Limits of determination; LOQ: Limits of quantification; RSD: Relative standard deviation; ANOVA: Analysis of variance.

Reversine inhibits MMP-1 and MMP-3 expressions by suppressing of ROS/MAPK/AP-1 activation in UV-stimulated human keratinocytes and dermal fibroblasts.

UVB has been shown to stimulate the generation of reactive oxygen species (ROS), which subsequently results in the activation of various intracellular signalling pathways and transcription factors (AP-1, NF-κB). These transcription factors are regulated by MAPKs, which increase cytokine and MMP expression. We examined the preventive effects of reversine on MMP-1 and MMP-3 expressions in NHEKs and NHDFs exposed to UVB irradiation. Also, we confirmed that reversine decreased pro-inflammatory cytokine expression in NHEKs. The mechanism underlying the MMP inhibitory effects of reversine occurred via the suppression of UVB-induced ROS generation and MAPK/AP-1 activation. Therefore, reversine is an effective therapeutic candidate for preventing skin photoageing.

Effects of Banxia Xiexin Decoction () on Cisplatin-Induced Apoptosis of Human A549 Lung Cancer Cells.

OBJECTIVE: To examinie the synergistic effects of Banxia Xiexin Decoction (, Known as Banhasasim-tang in Korean) extract (BXDE) on cisplatin-induced cytotoxicity in the A549 human lung cancer cell lines. METHODS: A549 cells were treated with varying concentrations (50-200 µg/mL) of cisplatin and BXDE alone or in combination for 96 h. We used 1-(4,5-dimethylthiazol-2-yl)-3,5-diphenylformazan assay and flow cytometry to analyze cell viability and apoptosis, respectively. RESULTS: The exposure of cells to cisplatin and BXDE alone or in combination decreased cell viability dose- and time-dependently (P<0.05), which was found to be mediated by the apoptotic pathway as confirmed by the increase in the annexin V+/propidium iodide- stained cell population and a ladder pattern of discontinuous DNA fragments. Furthermore, the apoptosis was inhibited by the pan-caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp (OMe) fluoromethylketone (z-VAD-FMK). CONCLUSIONS: BXDE significantly potentiated apoptotic effects of cisplatin in A549 cells. Moreover, apoptosis induced by BXDE might be the pivotal mechanism mediating its chemopreventative action against cancer.

Crotonis Fructus Extract Inhibits 12-O-Tetradecanoylphorbol-13-Acetate-Induced Expression of Matrix Metalloproteinase-9 via the Activator Protein-1 Pathway in MCF-7 Cells.

PURPOSE: Metastatic cancers spread from the primary site of origin to other parts of the body. Matrix metalloproteinase-9 (MMP-9) is essential in metastatic cancers owing to its major role in cancer cell invasion. Crotonis fructus (CF), the mature fruits of Croton tiglium L., have been used for the treatment of gastrointestinal disturbance in Asia. In this study, the effect of the ethanol extract of CF (CFE) on MMP-9 activity and the invasion of 12-O-tetradecanoylphorbol-13-acetate (TPA)-treated MCF-7 cells was examined. METHODS: The cell viability was evaluated using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. The expression of MMP-9 was examined by Western blotting, zymography, and real-time polymerase chain reaction. An electrophoretic mobility gel shift assay was performed to detect activator protein-1 (AP-1) DNA binding activity and cell invasiveness was measured by an in vitro Matrigel invasion assay. RESULTS: CFE significantly suppressed MMP-9 expression and activation in a dose-dependent manner. Furthermore, CFE attenuated the TPA-induced activation of AP-1. CONCLUSION: The results indicated that the inhibitory effects of CFE against TPA-induced MMP-9 expression and MCF-7 cell invasion were dependent on the protein kinase C δ/p38/c-Jun N-terminal kinase/AP-1 pathway. Therefore, CFE could restrict breast cancer invasiveness owing to its ability to inhibit MMP-9 activity.

Salvia miltiorrhiza extract inhibits TPA-induced MMP-9 expression and invasion through the MAPK/AP-1 signaling pathway in human breast cancer MCF-7 cells.

Cancer cell invasion is crucial for metastasis. A major factor in the capacity of cancer cell invasion is the activation of matrix metalloproteinase-9 (MMP-9), which degrades the extracellular matrix. Salvia miltiorrhiza has been used as a promotion for blood circulation to remove blood stasis. Numerous previous studies have demonstrated that S. miltiorrhiza extracts (SME) decrease lipid levels and inhibit inflammation. However, the mechanism behind the effect of SME on breast cancer invasion has not been identified. The inhibitory effects of SME on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced MMP-9 expression were assessed using western blotting, reverse transcription-quantitative polymerase chain reaction and zymography assays. MMP-9 upstream signal proteins, including mitogen-activated protein kinases and activator protein 1 (AP-1) were also investigated. Cell invasion was assessed using a matrigel invasion assay. The present study demonstrated the inhibitory effects of the SME ethanol solution on MMP-9 expression and cell invasion in TPA-treated MCF-7 breast cancer cells. SME suppressed TPA-induced MMP-9 expression and MCF-7 cell invasion by blocking the transcriptional activation of AP-1. SME may possess therapeutic potential for inhibiting breast cancer cell invasiveness.

Peucedanum japonicum Thunb. ethanol extract suppresses RANKL-mediated osteoclastogenesis.

The constituents of Peucedanum japonicum Thunb. (PJ) exhibit biological and pharmacological activities, including anti-obesity, anti-oxidant and anti-allergic activities. The aim of the present study was to examine in vitro effects of PJ in RANKL-induced signaling pathways, which determine osteoclast differentiation. PJ ethanol extract (PEE) exhibited anti-osteoporotic activity by disrupting the phospholipase C (PLC)-Ca2+-c-Fos/cAMP response element-binding protein (CREB)-nuclear factor of activated T cells, cytoplasmic 1 (NFATc1) signaling pathway during osteoclastogenesis. Murine bone marrow-derived macrophages (BMMs) were cultured and used to determine the effects of PJ in the receptor activator of nuclear factor κB ligand (RANKL)-mediated osteoclastogenesis. The effects of PEE in the RANKL-mediated signaling cascade were evaluated using a standard in vitro osteoclastogenesis system. PEE treatment of BMMs significantly reduced the number of RANKL-mediated tartrate resistant acid phosphatase (TRAP)-positive multinucleated cells (P<0.05 for 5 and 10 µg/ml PEE, P<0.01 for 25 and 50 µg/ml PEE), without cytotoxic effects. Furthermore, the expression of differentiation-related marker genes, including TRAP, Oscar, Cathepsin K, dendrocyte expressed seven transmembrane protein, ATPase H+ Transporting V0 Subunit D2 and NFATc1, were markedly suppressed. PEE induced a transient increase in free cytoplasmic Ca2+ ([Ca2+]i) mobilization via voltage-gated Ca2+ channels and PLC-sensitive pathways. Transient [Ca2+]i increase consequently resulted in the suppression of c-Fos, CREB and NFATc1 activities. These findings highlight the potential use of PJ in treating bone disorders caused by osteoclast overgrowth.

Sophorae Flos extract inhibits RANKL-induced osteoclast differentiation by suppressing the NF-κB/NFATc1 pathway in mouse bone marrow cells.

BACKGROUND: Sophorae Flos (SF) is a composite of flowers and buds of Styphnolobium japonicum (L.) Schott and has been used in traditional Korean and Chinese medicine for the treatment of hemostasis and inflammation. Previous studies reported that SF possesses anti-obesity properties, as well as anti-allergic, anti-proliferative, and anti-inflammatory activities. However, the effect of SF in bone resorption has not been studies. In this study, we examined the potential of SF extract (SFE) to inhibit receptor activator of NF-κB ligand (RANKL) -induced osteoclast differentiation in cultured mouse-derived bone marrow macrophages (BMMs). METHODS: BMMs, that act as osteoclast precursors, were cultured with M-CSF (50 ng/ml) and RANKL (100 ng/ml) for 4 days to generate osteoclasts. Osteoclast differentiation was measured by tartrate-resistant acidic phosphatase (TRAP) staining and the TRAP solution assay. Osteoclast differentiation marker genes were analyzed by the quantitative real-time polymerase chain reaction analysis. RANKLs signaling pathways were confirmed through western blotting. RESULTS: SFE significantly decreased osteoclast differentiation in a dose-dependent manner. SFE inhibited RANKL-induced osteoclastogenesis by suppressing NF-κB activation. By contrast, SFE did not affect phospholipase C gamma 2 or subsequent cAMP response element binding activation. SFE inhibited the RANKL-induced expression of nuclear factor of activated T cells c1 (NFATc1). CONCLUSIONS: SFE attenuated the RANKL-mediated induction of NF-κB through inhibition of IκBα phosphorylation, which contributed to inhibiting of RANKL-induced osteoclast differentiation through downregulation of NFATc1.

Identification and monitoring of Korean medicines derived from Cinnamomum spp. by using ITS and DNA marker.

In this study, we identified and evaluated the genetic relationships among Cinnamomum plants, which are used in traditional medicine. We also attempted to monitor the distribution of traditional medicines derived from Cinnamomum cassia by using DNA barcoding and a species-specific DNA marker. Plants of the genus Cinnamomum, and in particular C. cassia, are commonly used as medicinal herbs in the form of Cinnamomi Ramulus, Cinnamomi Cortex, and Cassiae Cortex Interior. However, it is difficult to distinguish among different Cinnamomum species based on morphological features, and so to overcome this limitation, nucleotide sequences of the internal transcribed spacer (ITS) region of Cinnamomum DNA were determined and compared. On the basis of the discrepancy in determined ITS sequences, a 408-bp product, amplified by the primer pair CC F1/CC R3, was developed as a C. cassia-specific DNA marker. Using the developed DNA marker in combination with the ITS 2 nucleotide sequence, we monitored imported and commercially supplied medicinal products derived from Cinnamomum plants in markets in Korean, China, and Japan. The results revealed that most of the specimens monitored were derived from C. cassia.

Influence of herbal combinations on the extraction efficiencies of chemical compounds from Cinnamomum cassia, Paeonia lactiflora, and Glycyrrhiza uralensis, the herbal components of Gyeji-tang, evaluated by HPLC method.

During decoction process, the ingredients of herbal formula interact with each other, such that therapeutic properties and chemical extraction characteristics are altered. The crude drugs, Cinnamomum cassia (CC), Paeonia lactiflora (PL), and Glycyrrhiza uralensis (GU), are the main herbal constituents of Gyeji-tang, a traditional herbal formula. To evaluate the chemical interaction between CC, PL, and GU during the course of decoction, quantification of 16 marker compounds in the herbal decoction, performed using a Box-Behnken experimental design, was carried out by HPLC-diode array detection using validated method. Correlations between the amounts of marker compounds from CC, PL, and GU were assessed by multiple regression analysis. The results obtained showed that amounts of single herb marker compounds significantly changed (usually decreased) by decoction in the presence of other herbs and that these changes depended on the chemical natures of the markers and the herbal medicines present. Results also demonstrated that the extraction efficiencies of marker compounds increased when the proportion of the herb containing them was increased and decreased in proportion to amounts of herbs added. In conclusion, chemical interactions between compositional herbal medicines may occur when herbs are co-decocted. This study provides insight of understanding the herbal interactions in herbal formulae.