Anti-Inflammatory Peroxidized Chlorahololide-Type Dimers Are Artifacts of Shizukaol-Type Dimers: From Phenomena Discovery and Confirmation to Potential Underlying Mechanism.

In our research on naturally occurring sesquiterpenes, eight shizukaol-type dimers, one chlorahololide-type dimer, and one sarcanolide-type dimer were isolated from the roots of Chloranthus fortunei. As the project was implemented, we accidentally discovered that shizukaol-type dimers can be converted into peroxidized chlorahololide-type dimers. This potential change was discovered after simulations of the changes in corresponding shizukaols showed that three peroxide products were generated (1-3), indicating that peroxidation reactions occurred. HPLC-HR-MS analysis results obtained for the shizukaol derivatives further demonstrate that the reaction occurred, and the type of substituent of small organic ester moieties at positions C-15' and C-13' of unit B were not decisively related to the reaction. Quantum chemical calculations of the mode dimer further demonstrated this phenomenon. The highest occupied molecular orbital (HOMO)-lowest unoccupied molecular orbital (LUMO) energy of the precursor and production revealed the advantageous yield of 4ß-hydroperoxyl production. Additionally, the potential reaction mechanism was speculated and validated using the free energy in the reaction which successfully explained the feasibility of the reaction. Finally, the anti-inflammatory activity of the precursors and products was evaluated, and the products of peroxidation showed better anti-inflammatory activity.

The intramolecular SN2 reaction tautomeric ent-Kauranoids isolated from the aerial parts of Isodon amethystoides.

As our ongoing searching for the bioactive natural terpenoids, nine ent-kauranoids (1-9), including three previously undescribed ones (1, 2, and 9), were isolated from the aerial parts of Isodon amethystoides. Their structures were elucidated on the basis of spectroscopic data analysis, including NMR, MS, and ECD. Compounds 1 and 2 were a pair of tautomeric compounds, which was confirmed by the HPLC analysis and low temperature NMR testing. The underlying mechanism of the tautomer was proposed as an intramolecular SN2 reaction, which was explained by quantum chemical calculation. The HOMO-LUMO gap and the free energy revealed the spontaneous of the tautomeric of the 1 and 2. Additionally, the similar phenomena were also found in the two groups of known compounds 3 and 4 and 6 and 7, respectively. Apart from the tautomer, compounds 3 and 4 can be hydrolyzed into 5 through ester hydrolysis in CDCl3, while compounds 6, 7 can be hydrolyzed into 8 through ester hydrolysis. These phenomena were also confirmed through HPLC analysis and low temperature nuclear magnetic resonance tests and the mechanism was studied using quantum chemical calculation.

Structure-diversified terpenoids from Salvia prattii and their protective activity against alcoholic liver diseases.

Eleven previously unreported compounds (1-11), including five diterpenoids (1-5) and six sesquiterpenoids (6-11), together with two known diterpenoids (12-13), have been isolated from the roots of Salvia prattii. Their structures were comprehensively elucidated through spectroscopic methods, and their configurations were established using computational 13C nuclear magnetic resonance and electronic circular dichroism. Compound 1 was found to be an abietane-type diterpenoid with a novel rearrangement generated from the cleavage of the C-4/5 chemical bond, 20-methyl shift, and the rearrangement of the C-10 side chain. Compounds 2-3 were the third and fourth examples of arrangement seco-norabietanes with a spiro-lactone ring. We evaluated all compounds for their protective effects against alcoholic liver diseases (ALD). Compound 2 exhibited potential protective activity and hence can be used as a novel anti-ALD candidate.

Sesquiterpene dimers from Chloranthus fortunei and their protection activity against acute lung injury.

Chloranthus fortunei (family Chloranthaceae), a perennial herb, widely distributed in south China with an altitude of 170-340 m. The whole plants were used as an anti-inflammatory agent for the treatment of cough, arthritis and tumor. Five previously unreported compounds fortulactones A-E were isolated from the aerial part of Chloranthus fortunei. Their structures were elucidated using 1D/2D NMR and HRESIMS and their absolute configuration were determined using the ECD excitron chirality method. All isolates were tested for inhibitory effects on the NO production of liposaccharide (LPS)-induced RAW 264.7 macropahges. The most potent compound 1 was further evaluated its protective activity against LPS stimulated A549 cells, the ELISA kits results showed the abnormal states of MDA and SOD were corrected to a certain extent. Meanwhile, the pro-inflammatory cytokine, such as TNF-α, IL-6 and IL-1ß were also attenuated. In conclusion, these results showed that 1 exhibited therapeutic potential for ameliorating ALI.

[Impacts of multicomponent environment on intrinsic dissolution rate of berberine in biopharmaceutics classification system of Chinese materia medica].

To illustrate the intrinsic dissolution rate(IDR) involved in biopharmaceutics classification system of Chinese materia medica(CMMBCS), investigate the effect of artificial multicomponent environment on IDR of berberine, by using a non-disintegrating disk, and summarize related rules by using the obtained data. Progressive levels design was used to investigate the effects of single component environment (puerarin, baicalin, and glycyrrhizic acid); double-component environment(puerarin+baicalin, puerarin+ glycyrrhizic acid, baicalin+glycyrrhizic acid); and triple-component environment(puerarin+baicalin+glycyrrhizic acid) on IDR of berberine, laying a foundation for further researches on the absorption mechanism of multiple components.

[Study on biopharmaceutics classification system for Chinese materia medica of extract of Huanglian].

One of the advantages of biopharmaceutics classification system of Chinese materia medica (CMMBCS) is expanding the classification research level from single ingredient to multi-components of Chinese herb, and from multi-components research to holistic research of the Chinese materia medica. In present paper, the alkaloids of extract of huanglian were chosen as the main research object to explore their change rules in solubility and intestinal permeability of single-component and multi-components, and to determine the biopharmaceutical classification of extract of Huanglian from holistic level. The typical shake-flask method and HPLC were used to detect the solubility of single ingredient of alkaloids from extract of huanglian. The quantitative research of alkaloids in intestinal absorption was measured in single-pass intestinal perfusion experiment while permeability coefficient of extract of huanglian was calculated by self-defined weight coefficient method.

[Overall intestinal permeability of multiple components in lotus leaf by in situ single pass intestinal perfusion models].

To investigate the overall intestinal permeability of multiple components in lotus leaves and make clear the interaction in composition absorption process. Rat single-pass intestinal perfusion technique was used, and the results showed that the Peff values of nuciferine, demethylanuciferine, rutin, quercetin, kaempferol from lotus leaf were greater than 0.5×10⁻4 cm•s⁻¹. In the biopharmaceutics classification system (BCS) intestinal permeability property, these ingredients were high permeable components, while the hyperin was low permeable component. However, in the multi-component environment of the lotus leaf extract, component permeation was changed. Semi quantitative analysis of the unclear components showed that under the multi-component environment, four in seven components with relatively high contents had a Peff value less than 0.5×10⁻4 cm•s⁻¹, indicating these 4 components were of low permeability, while other 3 components were of high permeability. The results could be valuable to make clear the overall intestinal permeability of multiple components in lotus leaf, and lay a foundation for studying the mechanism of the lipid-lowering effect of lotus leaf.

[Validation of in situ single pass perfusion model based on P-gp].

To validate in situ rats intestinal single pass perfusion model based on P-glycoprotein (P-gp). Firstly, phenol red perfusion was carried out to verify the close connection structure of intestinal epithelial cells, and the integrity of the intestinal epithelium, with a gravimetric method for correcting water flux. The level of phenol red was determined by high performance liquid chromatography (HPLC) both before and after perfusion. Secondly, the positive drug digoxin specified by FDA was used to validate the model. After different mass concentrations of verapamil were given in the rats, the absorption parameters of digoxin in ileum of rats were observed and compared. The results showed that the phenol red was absorbed in rats ileum segment, with an effective permeability coefficient of (1.09±0.62)×10 ⁻6 cm•s ⁻¹. The experiment results indicated that the close connection structure of intestinal epithelial cells was normal, and the integrity of the intestinal epithelium was maintained well. In digoxin perfusion experiment, in case no verapamil was given, digoxin showed certain degree of absorption in rat ileum, with an effective permeability coefficient (Peff) of (1.07±0.59)×10 ⁻5 cm•s ⁻¹; after mass concentrations of 0.01,0.1 mmol•L ⁻¹ verapamil were given, the absorption of digoxin was on the rise in rat ileum, with an effective permeability coefficient Peff of (1.58±0.69)×10 ⁻5, (3.28±0.95)×10 ⁻5 cm•s ⁻¹ respectively (P<0.05). Digoxin perfusion experiment verified that P-gp expression in small intestine epithelium was intact and can be used in the research of P-gp efflux transporter.

[Overall intestinal permeability of multiple components in lotus leaves].

In the study of biopharmaceutics classification system of Chinese materia medica (CMMBCS), the interactions of multiple components in the absorption should be taken into consideration in simultaneous multi-component determination. To investigate the absorption of multiple components, the in vitro everted gut sac model was used in this study, wtih lotus leaves as the research object. Aquantitative analysis was also carried out for the known components in this study. Totally 19 components in lotus extracts were absorbed by the intestinal tract, the Papp levels of the known components were nuciferine (1×10⁻5-1×10⁻6 cm•s⁻¹), rutin (1×10⁻6-1×10⁻7 cm•s⁻¹), hyperoside (1×10⁻6 cm•s⁻¹), isoquercitrin (1×10⁻6-1×10⁻7 cm•s⁻¹) and astragalin (1×10⁻6-1×10⁻7 cm•s⁻¹), respectively. These components showed a low permeability under a multi-component environment. This study was carried out to lay a foundation for further relevant target studies for different categories of components.

[Study on property of biopharmaceutics classification system of berberine in Huanglian decoction].

The study of single component in the multicomponent environment is one of the basic researches for biopharmaceutics classification system of Chinese materia medica (CMMBCS). That is to say, the classification research shall be based on the respective lift of solubility and permeability in the multicomponent environment, besides solubility and intestinal permeability of the single component. We chose berberine as the main research object to investigate the changes of its solubility and intestinal permeability in Huanglian decoction. Shake-flask and HPLC were used to detect the solubility of berberine in different pH buffer solutions and different concentrations of Huanglian decoction. In situ single-pass intestinal perfusion (SPIP) and intestinal perfusion with venous sampling (IPVS) were carried out to study berberine's intestinal absorption and absorption into blood, respectively.