Discovery and validation of COX2 as a target of flavonoids in Apocyni Veneti Folium: Implications for the treatment of liver injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Apocyni Veneti Folium (AVF), a popular traditional Chinese medicine (TCM), is known for its effects in soothing the liver and nerves and eliminating heat and water. It is relevant from an ethnopharmacological perspective. Pharmacological research has confirmed its benefits on antihypertension, antihyperlipidemia, antidepression, liver protection, immune system boosting, antiaging, and diabetic vascular lesions. Previous studies have shown that flavonoids, the active ingredients, have a hepatoprotective effect. However, the exact mechanism has not been clarified. AIM OF THE STUDY: This study aimed to identify the active flavonoids in AVF and their corresponding targets for liver injury. Multiple methods were introduced to confirm the targets. MATERIAL AND METHODS: AVF compounds were analyzed using liquid chromatography-mass spectrometry (LC-MS). Then, network pharmacology was utilized to screen potential hepatoprotection targets of the compounds. An enzyme activity assay was performed to determine the effect of the compounds on the targets. Biolayer interferometry (BLI) was applied to confirm the direct interaction between the compounds and the targets. RESULTS: A total of 71 compounds were identified by LC-MS and 19 compounds and 112 shared targets were screened using network pharmacology. These common targets were primarily involved in the TNF signaling pathway, cancer pathways, hepatitis B, drug responses, and negative regulation of the apoptotic process. Flavonoids were the primary pharmacological substance basis of AVF. The cyclooxygenase 2 (COX2) protein was one of the direct targets of flavonoids in AVF. The enzyme activity assay and BLI-based intermolecular interactions demonstrated that the compounds astragalin, isoquercitrin, and hyperoside exhibited stronger inhibition of enzyme activity and a higher affinity with COX2 compared to epigallocatechin, quercetin, and catechin. CONCLUSIONS: COX2 was preliminarily identified as a target of flavonoids, and the mechanism of the hepatoprotective effect of AVF might be linked to flavonoids inhibiting the activity of COX2. The findings can establish the foundation for future research on the traditional hepatoprotective effect of AVF on the liver and for clinical studies on liver disorders.

A strategy for quality evaluation of salt-treated Apocyni Veneti Folium and discovery of efficacy-associated markers by fingerprint-activity relationship modeling.

In this study, a fingerprint-activity relationship between chemical fingerprints and hepatoprotective activity was established to evaluate the quality of salt-treated Apocyni Veneti Folium (AVF). Characteristic fingerprints of AVF samples exposed to different concentrations of salt were generated by ultrafast liquid chromatography tandem triple time-of-flight mass/mass spectrometry (UFLC-Triple TOF-MS/MS), and a similarity analysis was performed based on common characteristic peaks by hierarchical clustering analysis (HCA). Then, the hepatoprotective activity of AVF against CCl4-induced acute liver damage in mice was investigated by assessing biochemical markers and histopathology, which showed that a high dose of AVF exposed to low levels of salt stress produced a marked amelioration of hepatic damage compared with the other salt-treated AVF. Finally, fingerprint-activity relationship modeling, which was capable of discovering the bioactive markers used in the quality evaluation, was investigated by the chemical fingerprints and the hepatoprotective activities utilizing multivariate statistical analysis, gray correlation analysis (GCA) and bivariate correlation analysis (BCA). The results showed that the accumulation of polyphenols, such as flavonoids and phenolic acids, in AVF subjected to low levels of salt stress could result in the effective scavenging of free radicals. Therefore, the present study may provide a powerful strategy to holistically evaluate the quality of salt-treated AVF in combination with chemical fingerprint and bioactivity evaluation.

Simultaneous determination of 19 fatty acids in Antrodia camphorata by derivatized GC-MS and evaluation of antioxidant activity of Antrodia camphorata crude oil.

To analyze the components of the fatty oil extracted from Antrodia camphorata fungus powder and to evaluate the antioxidant activity of A. camphorata crude oil. A derivatized gas chromatography mass spectrometry (GC-MS) was developed for the quantification of 19 fatty acids in extracts of A. camphorata obtained by supercritical carbon dioxide (SC-CO2). Good separation was obtained under the optimized chromatographic conditions and 19 target compounds after methyl-esterification were identified by GC-MS on a HP-VOC capillary column (60 m × 320 µm × 1.8 µm) with an initial temperature set at 80 °C. The validity of the established method was examined experimentally with good linearity, intra-assay precisions, repeatability, stability and recovery. The antioxidant activity of A. camphorata crude oil was evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical-scavenging assay. Nineteen fatty acids showed good linearity over the tested ranges (r > 0.9956) and the recovery ranged from 93.47 to 104.89 %. The crude oil extracted from A. camphorata fungus powder also revealed its antioxidant activity. It was the report about simultaneous determination of 19 fatty acids in A. camphorata and antioxidant activity of its crude oil for the first time. The established method might also be utilized for the investigation of edible plant materials and agricultural products containing fatty acids.

Simultaneous determination of 10 nucleosides and nucleobases in Antrodia camphorata using QTRAP LC-MS/MS.

A liquid chromatography-triple-quadrupole linear ion trap mass spectrometry (LC-QTrap-MS) analysis has been developed for the identification and quantification of 10 nucleosides and nucleobases in extracts of Antrodia camphorata. The method was successfully used to qualitatively identify for six nucleosides namely, cytidine, uridine, inosine, guanosine, thymidine, adenosine and four nucleobases namely, uracil, guanine, xanthine, adenine in A. camphorata. Under optimized chromatographic conditions, good separation for 10 target compounds were obtained on an Agilent HC-C18(2) column (4.6 × 250 mm, 5 µm) eluted by a mobile phase of 5 mM ammonium acetate solution-methanol at a flow rate of 0.5 mL/min. Data acquisition was carried out in multiple reaction monitoring transition mode. Additional identification and confirmation of target compounds were performed using the enhanced product ion modus of the linear ion trap. It was the first report about simultaneous analysis of nucleosides and nucleobases in A. camphorata using this method. These results demonstrated that the QTRAP LC-MS/MS was a useful tool for quality evaluation of some medicinal plant products by using nucleosides and nucleobases as chemical markers. This method might also be utilized for the investigation of edible plant materials and agricultural products containing nucleosides and nucleobases.

[Preliminary analysis of quality of Antrodia camphorata powder].

OBJECTIVE: To proceed a preliminary analysis of the quality of Antrodia camphorata powder. METHODS: The contents of water-soluble extract were detected according to the standards stated in Chinese Pharmacopoeia. UV-VIS was used to analyze total polysaccharide and triterpenoid. HPLC method was applied for the analysis of adenosine in Antrodia. camphorata. Besides, volatile compounds were analyzed by HSGC-MS. RESULTS: The contents of water-soluble extract (37.26% - 40.98%), total polysaccharide (5.45% - 8.08%), total triterpenoid (2.44% - 2.87%) and adenosine (0.0470% - 0.0604%) were obtained respectively. 49 volatile compounds were identified in Antrodia camphorata powder. CONCLUSION: The established method can be used for the quality control of Antrodia camphorata powder.

[Study on water-soluble iron, heavy metals and harmful elements of Magnetitum].

OBJECTIVE: To measure the contents of the water-soluble iron, five heavy metals and harmful elements in Magnetiturn and provide a basis for the quality control and safety evaluation of Magnetitum. METHOD: Iron (Fe), lead (Pb), cadmium (Cd) and copper (Cu) were determined by atomic absorption spectrometry (AAS); arsenic (As) and mercury (Hg) were determined by atomic fluorescence spectrometry (AFS). RESULT: The mean content of element iron is 764.30 mg x kg(-1). The contents of five water-soluble heavy metals and harmful elements in Magnetitum were within the safety range. The recovery of the standard addition was in the range of 93.7% - 110.6%, and the RSD was less than 5.0%. CONCLUSION: Analyzing the water-soluble iron, heavy metals and harmful elements in Magnetitum is effective to the quality control and the safety evaluation of magnetitum.

Pentacyclic triterpenes from the resin of Liquidambar formosana.

Two new pentacyclic triterpenes and eight known pentacyclic triterpenes were isolated from the petroleum ether extract of Liquidambaris Resina. The structural elucidation of these compounds was determined by spectroscopic data interpretation. Their cytotoxic and antiplatelet aggregation activities were examined to find potent cytotoxic and antiplatelet aggregation compounds from natural resources. The results showed that 3-keto oleane triterpenes had strong cytotoxicity against MDA-MB-435S cancer cells and that 28-carboxyl oleane triterpenes possessed significant inhibition of platelet aggregation induced by ADP.