Comparison of Murraya exotica and Murraya paniculata by fingerprint analysis coupled with chemometrics and network pharmacology methods.

There are two source plants for the traditional Chinese medicine Murrayae Folium et Cacumen (MFC) in Chinese Pharmacopoeia, i.e. Murraya exotica L. and M. paniculata (L.) Jack. Herein, a chemical comparison of M. exotica and M. paniculata by high performance liquid chromatography (HPLC) fingerprint analysis coupled with chemometrics and network pharmacology was performed. The main peaks in the fingerprints were identified by liquid chromatography coupled with ion trap/time-of-flight mass spectrometry (LC-IT-TOF-MS) and authenticated by references. The chemometrics results showed that the HPLC fingerprints of these two species were clearly divided into two categories using hierarchical cluster analysis (HCA) and principal component analysis (PCA), and a total of 13 significantly differentiated markers were screened out by orthogonal partial least squares-discriminant analysis (OPLS-DA). However, the following network pharmacology analysis showed that these discriminated markers were found to act via many common targets and metabolic pathways, indicating the possibly similar pharmacological effects and mechanisms for M. exotica and M. paniculata. The above results provide valuable evidence for the equivalent use of these two plants in clinical settings. Moreover, the chromatographic fingerprint analysis coupled with chemometrics and network pharmacology supplies an efficient approach for the comparative analysis of multi-source TCMs like MFC.

The complete chloroplast genome of Melicope pteleifolia (Champ. ex Benth.) T. G. Hartley (Rutaceae).

Melicope pteleifolia, an important medicinal and horticultural plant, has antipyretic, anti-inflammatory, and analgesic effects. Here, the complete chloroplast genome of M. pteleifolia was sequenced and its phylogenetic relationship was investigated. The complete chloroplast genome of M. pteleifolia was 159,014 bp in size, including a pair of inverted repeat regions (IR, 27,640 bp), a large single copy region (LSC, 85,124 bp), and a small single copy region (SSC, 18,610 bp). The GC content of the chloroplast genome was 38.3%. A total of 133 genes were annotated, including 88 protein-coding genes, 8 rRNA genes, and 37 tRNA genes. Phylogenetic analysis revealed that M. pteleifolia clusters together with species of Toddalia, Zanthoxylum, Tetradium, Phellodendron, and Casimiroa.

[Phenolic constituents from stems of Ilex asprella].

Phytochemical investigation on Ilex asprella stems by using various chromatographic techniques led to the isolation of 18 phenolic constituents. Based on spectroscopic data analyses and/or comparison of the spectroscopic data with those in literature, these constituents were identified, including two lignans (1, 2), five phenylpropanes (3-7), six chlorogenic analogues (8-13), and five benzoic analogues (14-18). Among them, compounds 3-7, 9, 11, 13, 14, 17, and 18 were isolated from genus Ilex for the first time, and 2, 8, 10, 15, and 16 were isolated from this species for the first time. The in vitro anti-inflammatory assay results showed that compounds 8, 9, 11, 13, and 15 possessed moderate inhibition on the NO production in RAW264.7 cells with IC50 values of 51.1-85.8 µmol·L⁻¹. The present study brought preliminary reference for the clarification of therapeutic ingredients of I. asprella with anti-inflammatory efficacy and its quality evaluation.

Optimization of Storage Conditions of the Medicinal Herb Ilex asprella against the Sterigmatocystin Producer Aspergillus versicolor Using Response Surface Methodology.

The root of Ilex asprella is a commonly used herb in Southern China, and also constitutes the main raw material of Canton herbal tea. I. asprella is readily contaminated by mildew because of rich nutrients. Aspergillus versicolor producing sterigmatocystin is one of the most common molds that contaminate foodstuffs and medicinal herbs. Previous study on the evaluation of fungal contamination on samples of I. asprella revealed that A. versicolor was the dominant contaminant. In this study, experiments based on response surface methodology combined with central composite design were carried out to determine the optimal storage conditions of I. asprella to minimize the contamination of sterigmatocystin. The herb, manually innoculated with A. versicolor, was stored under different temperatures (20⁻40 °C) and humidity (80⁻95%) for seven days. The effects of temperature and humidity were evaluated using total saponin, polysaccharide and the sterigmatocystin levels as quality indexes. The results showed that A. versicolor grew quickly and produced large amounts of sterigmatocystin on I. asprella, at humidity ranging from 85% to 90% and temperatures above 26 °C. Meanwhile, total saponin and polysaccharide amounts were reduced significantly. These findings suggested that I. asprella samples should be stored in an environment with humidity and temperature below 85% and 26 °C, respectively, to reduce A. versicolor growth and sterigmatocystin production.

Asprenols A-H, phenolic constituents from the stems of Ilex asprella.

Encouraged by the in vivo anti-inflammatory effect of aqueous extract of Ilex asprella stems, a further phytochemical investigation on I. asprella stems oriented by the in vitro NO production inhibition in RAW264.7 cells was conducted, which led to the isolation of eight new phenolic constituents, namely asprenols A-H (1-8), together with 12 known ones (9-20). The structures of the new compounds were established by extensive spectroscopic data analyses of HR-ESI-MS, IR, UV, and 1D and 2D NMR, and the absolute configurations were determined by comparison of experimental and calculated ECD analyses. All isolated were evaluated for the inhibition against NO production in RAW 264.7 cells, and several compounds showed moderate inhibitory effect.

Ilex asprella aqueous extracts exert in vivo anti-inflammatory effects by regulating the NF-κB, JAK2/STAT3, and MAPK signaling pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Ilex asprella (Hook. et Arn.) Champ. ex Benth. (IA) is a representative medicinal plant from the South of the Five Ridges of China. Its roots (RIA) and stems (SIA) have been traditionally used for the inflammation-related diseases, such as acute and chronic pharyngitis, cough, and sore throats. AIM OF THE STUDY: To evaluate the in vivo anti-inflammatory effects of IA extracts to provide evidence for its traditional use and to enhance the knowledge of the medicinal properties of IA. MATERIALS AND METHODS: Models of xylene-induced ear edema in mice and carrageenan-induced paw edema in rats were used for the pharmacological evaluations. The mice were randomly divided into eight groups (n = 10 per group): a model group, a positive control group [dexamethasone (Dex), 10 mg/kg, intragastrically (i.g.)], RIA aqueous extract groups with three dosages (30, 15, and 7.5 mg/kg, i.g.), and SIA aqueous extract groups with three dosages (60, 30, and 15 mg/kg, i.g.). The rats were randomly divided into eight groups (n = 6 per group): a model group, a positive control group [acetylsalicylic acid (ASA), 300 mg/kg, i.g.], RIA groups with three dosages (80, 40, and 20 mg/kg, i.g.) and SIA aqueous extract groups with three dosages (160, 80, and 40 mg/kg, i.g.). Histological examinations of the ear and paw tissues were observed by hematoxylin-eosin (HE) staining, and neutrophil elastase levels were assessed in ear tissues by immunohistochemical analysis. Serum levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1ß (IL-1ß) were measured by ELISAs, and expression levels of TNF-α, IL-6, and IL-1ß in rat paw tissues were measured by RT-PCR. The signal transduction proteins p65, IκBα, Janus kinase 2 (JAK2), signal transducer and activator of transcription 3 (STAT3), p38 mitogen-activated protein kinases (p38), extracellular regulated protein kinases (ERK1/2), and c-Jun N-terminal kinase (JNK) in the rat paw tissues were investigated by western blot analyses. RIA and SIA were characterized by HPLC and LC-MS analyses, and the components were confirmed by comparison with isolated compounds. RESULTS: Intragastric administration with RIA (30 mg/kg) and SIA (60, 30 mg/kg) significantly mitigated ear edema in mice. RIA administration at 80 and 40 mg/kg reduced paw edema in rats 2‒3 h after injection. SIA administration with 160 mg/kg inhibited paw edema in rats after the injection of carrageenan for 1‒4 h, and SIA administration at 80 mg/kg inhibited paw edema after the injection of carrageenan for 2‒4 h. Meanwhile, RIA (80, 40 mg/kg) and SIA (160, 80 mg/kg) reduced inflammatory cell infiltration in the ear and paw tissues and infiltration of neutrophil leukocytes in rat paw tissues. RIA (80, 40, and 20 mg/kg) and SIA (160, 80, and 40 mg/kg) notably inhibited the increases of TNF-α, IL-6 and IL-1ß in the serum and mRNA expression in the rat paw tissues. RIA (80, 40 mg/kg) and SIA (160, 80 mg/kg) reduced the p-p65/p-IκBα, p-JAK2/p-STAT3, and p-p38/p-ERK1/2/p-JNK levels in the pathological tissues of the animals. Phenolic acids and triterpenoids likely contributed to the anti-inflammatory activity. CONCLUSIONS: Both RIA and SIA aqueous extracts showed anti-inflammatory effects in vivo in a dose-independent manner (20‒80 and 40‒160 mg/kg, respectively). The underlying mechanisms are mediated by inhibiting the pro-inflammatory cytokines TNF-α, IL-6, and IL-1ß via regulation of the NF-κB, JAK2/STAT3, and MAPK signaling pathways. The present results provided pharmacological evidence that stems are alternative medicinal parts of IA but function at different doses. Additionally, this study supports the use of IA as an anti-inflammatory herbal medicine.

Three new triterpenoid saponins from the stems of Ilex asprella.

Three new triterpenoid saponins, namely asprellinoids A-C (1-3), featuring a sulfate substitution in sugar moiety, were isolated from the stems of Ilex asprella (Hook. et Arn.) Champ. ex Benth. Their structures were elucidated by the spectroscopic data analyses including HR-ESI-MS, IR, and NMR spectra, and chemical method.

[Chemical constituents from stems of Ilex asprella].

Phytochemical investigation on the stems of Ilex asprella by using various chromatographic techniques led to the isolation of 13 compounds. By spectroscopic analyses and comparisons the spectral data with those in literatures, these compounds were identified as salicifoneoliganol(1), rel-(7R,8S)-3,3',5-trimethoxy-4',7-epoxy-8,5'-neolignan-4,9,9'-triol 9-ß-D-glucopyranoside(2),(+)-cycloolivil(3),(+)-syringaresinol-4'-O-ß-D-monoglucoside(4), liriodendrin(5), caffeic acid (6), 3,4-dihydroxy-5-methoxybenzaldehyde(7), benzene-1,2,4-triol(8), 3,4,5-trimethoxyphenyl-1-O-ß-D-apiofuranosyl(1″→6')-glucopyranoside(9), aeculetin(10), cryptochlorogenic acid ethyl ester(11), chlorogenic acid ethyl ester(12), and rel-5-(3S,8S-dihydroxy-1R,5S-dimethyl-7-oxa-6-oxobicyclo [3,2,1]oct-8-yl)-3-methyl-2Z,4E-pentadienoic acid(13). Among them, compounds 7, 8, 11, and 13 were isolated from genus Ilex for the first time, and 1-3, 9, 10, and 12 were isolated from this speciesfor the first time. The anti-inflammatory assay results of these compounds showed that compounds 1 and 9 showed moderate inhibitory effect against NO production in RAW 267. 4 cells with IC50 values of 35.7 and 50.6 µmol•L⁻¹, in vitro respectively, whereas compound 10 showed weak inhibition(IC50 value 98.7 µmol•L⁻¹).

Prenylated benzoic acid derivatives from the stem of Euodia lepta.

Two new prenylated benzoic acid derivatives, leptoic acid A and (+)-S-anodendroic acid (1-2), along with one known compound, 2,2-dimethyl-2H-1-benzopyran-6-carboxylic acid (3) were isolated from the stem of Euodia lepta (spreng.) Merr. Their structures were elucidated on the basis of the chemical and spectroscopic evidence.