Total triterpenes of Euphorbium alleviates rheumatoid arthritis via Nrf2/HO-1/GPX4 pathway / 中国中药杂志

This study aims to investigate the effect and mechanism of total triterpenes of Euphorbium in treating rheumatoid arthritis(RA). The rat model of RA was established with Freund's complete adjuvant(FCA). Male rats were randomly assigned into control, model, Tripterygium glycosides(7.5 mg·kg~(-1)), and low-, medium-, and high-dose total triterpenes of Euphorbium(32, 64, and 128 mg·kg~(-1), respectively) groups, with 10 rats in each group. In other groups except the control group, 0.2 mL FCA was injected into the right hind toe. Rats in the intervention groups were administrated with corresponding drugs by gavage, and the control group and the model group with the same volume of 0.5% CMC-Na solution once a day. During the treatment period, the swelling degree of the hind paw was measured and the arthritis was scored until day 30. At the end of drug administration, the pathological changes of the joint tissue were observed by hematoxylin-eosin staining. The content of malondialdehyde(MDA), glutathione(GSH), and Fe~(2+) and the activity of superoxide dismutase(SOD) in the joint tissue were measured by biochemical colorimetry. RT-PCR was performed to determine the mRNA levels of nuclear factor erythroid 2-related factor 2(Nrf2), glutathione peroxidase 4(GPX4), and acyl-CoA synthetase long chain family member 4(ACSL4) in the joint tissue. Western blot was employed to determine the protein levels of Nrf2, Kelch-like ECH-associated protein 1(Keap1), heme oxygenase-1(HO-1), NAD(P)H quinone oxidoreductase 1(NQO1), SOD2, GPX4, and ACSL4 in the joint tissue. The results showed that the treatment with Tripterygium glycosides(7.5 mg·kg~(-1)) and total triterpenes of Euphorbium(32, 64, and 128 mg·kg~(-1)) alleviated the swelling degree of bilateral hind limbs, decreased the arthritis score, reduced joint tissue lesions and the content of MDA and Fe~(2+) in the joint tissue, and increased GSH content and SOD activity. Furthermore, the interventions up-regulated the protein and mRNA levels of Nrf2 and GPX4, down-regulated the protein and mRNA levels of ACSL4, and up-regulated the protein levels of Keap1, NQO1, HO-1, and SOD2 in the Nrf2/HO-1/GPX4. In summary, the total triterpenes of Euphorbium can treat RA by inhibiting lipid peroxidation and abnormal ferroptosis, which may involve the Nrf2/HO-1/GPX4 signaling pathway.

Investigation on in Vitro Antioxidant Activity and Chemical Composition of Different Polar Parts of Extract of Rosa cymosa Roots / 中国实验方剂学杂志

Objective:To investigate the antioxidant activity and chemical composition of 75% ethanol extract of <italic>Rosa cymosa</italic> roots and its different polar parts. Method:The 75% ethanol extract of <italic>R. cymosa</italic> roots was divided into dichloromethane， ethyl acetate， <italic>n</italic>-butanol and water parts by organic solvent extraction. <italic>In vitro</italic> antioxidant activity of each fraction was determined by 1，1-diphenyl-2-picrylhydrazyl （DPPH） and 2，2'-azino-bis（3-ethylbenzothiazoline-6-sulfonic acid） （ABTS） free radical scavenging assays， as well as ferric reducing antioxidant power （FRAP） test. The contents of total triterpenes， total phenols， total tannins and condensed tannins in each fraction were determined by spectrophotometry. SPSS 24.0 software was used to conduct Pearson correlation analysis between the antioxidant activity of each fraction and the content of the main components， and then the main active fraction and the main active components were determined. The chemical constituents of the active fraction was analyzed by ultra performance liquid chromatography-quadrupole-time-of-flight mass spectrometry （UPLC-Q-TOF-MS/MS）， and the structures of the main chromatographic peaks were predicted. Result:Each fraction of <italic>R. cymosa</italic> roots had certain antioxidant activity， and there was a significant dose-effect relationship within a certain concentration range， but the antioxidant activity of different polar parts was different. In DPPH and ABTS free radical scavenging tests， the antioxidant activity of each fraction and vitamin C （VC， the positive drug） was ranked as ethyl acetate fraction>VC><italic>n</italic>-butanol fraction>ethanol extract>water fraction>dichloromethane fraction. In FRAP test， the activity of ethyl acetate fraction was weaker than that of VC， and the other order was unchanged. The contents of total triterpenes， total phenols， total tannins and condensed tannins in ethyl acetate fraction were 3.81%， 50.33%， 3.32%， and 39.79%， in <italic>n</italic>-butanol fraction were 0.88%， 41.42%， 2.25% and 23.55%， in ethanol extract were 2.90%， 41.95%， 3.43% and 20.14%， in water fraction were 0， 26.80%， 16.90% and 7.57%， and in dichloromethane fraction were 21.23%， 12.90%， 1.59%， and 6.17%， respectively. Correlation analysis results showed that the contents of total phenols and condensed tannins were positively correlated with the antioxidant activity， the contents of total triterpenes were negatively correlated with the antioxidant activity， and the correlation between total tannins and antioxidant activity was not obvious. A total of 26 compounds were identified from the ethyl acetate fraction by UPLC-Q-TOF-MS/MS， including 11 condensed tannins， 4 hydrolysable tannins， 6 triterpenes， 3 flavonoids， 1 benzoic acid derivative and 1 chlorogenic acid analogue. Conclusion:Ethyl acetate fraction is the main antioxidant active site of <italic>R. cymosa</italic> roots， and phenols mainly composed of condensed tannins are the main active components. The results can provide experimental basis for the development of natural antioxidants.

Establishment of HPLC Fingerprint ，Cluster Analysis ，Principal Component Analysis and Content Determination of Paliurus ramosissimus Total Triterpenes / 中国药房

OBJECTIVE：To establish fingerprint of Paliurus ramosissimus total triterpenes ，and to conduct cluster analysis and principal component analysis ，and to determine the content of the main component paliurusene. METHODS ：HPLC method was adopted. The determination was performed on Agilent PheHex column with mobile phase consisted of methanol- 0.05％ phosphoric acid solution （gradient eluetion ） at the flow rate of 1 mL/min. The detection wavelength was set at 320 nm，and column temperature was 30 ℃. The sample size was 5 μL. Using paliurusene as reference，HPLC fingerprints of 10 batches of P. ramosissimus total triterpenes were drawn. Similarity evaluation was performed by using TCM Chromatographic Fingerprint Similarity Evaluation System （2012 edition），and the common peaks were confirmed. Cluster analysis and principle component analysis were performed by using SPSS 26.0 software. The content of paliurusene was determined by same HPLC method. RESULTS：There were totally 6 common peaks in HPLC fingerprint of 10 batches of P. ramosissimus total triterpenes. The similarity was more than 0.990；one of six common peaks was identified as paliurusene. The results of cluster analysis showed that 10 batches of samples could be clustered into 4 categories，including S 1，S2，S3-S6 and S 7-S10. The results of principal component analysis showed that the accumulative variance contribution rate of primary 2 principal components was 99.430％. Comprehensive score ranking was S 1＞S9＞S8＞S7＞S10＞S2＞S3＞S5＞S6＞S4. The linear range of paliurusene concentration was 33.7-844.0 μg/mL（r＝0.999 9）. RSDs of precision ，reproducibility and stability （24 h）tests were all lower than 2％. Average recovery was 99.75％（RSD＝1.13％，n＝6）. The average contents of paliurusene in 10 batches of P. ramosissimus total triterpenes was 0.576％-0.712％. CONCLUSIONS ：Established HPLC fingerprint and content d etermination method are reliable and stable ， and can be used for the quality control of P. ramosissimus .

Optimization of Extraction Technology of Total Triterpenes from the Leaves of Cornus officinalis by Box-Behnken Design-response Surface Methodology / 中国药房

OBJECTIVE：To optimize the extraction technology for total triterpenes from the leaves of Cornus officinalis . METHODS：Based on the full swelling of the leaves of C. officinalis ，total triterpenes was extracted with heating reflux method. The effects of ethanol concentration ，liquid-solid ratio ，extraction time and extraction times on the contents of total triterpenes from the leaves of C. officinalis were investigated by single factor test. Using oleanolic acid as control ，the contents of total triterpenes were detected by UV spectrometry. On the basis of single factor test ，fixing the times of extraction a s 3 times，taking the contents of total triterpenes as response value ，using ethanol volume fraction ，solid-liquid ratio and extraction time as factors ， Box-Behnken design-response methodology was used to optimize the extraction technology of total triterpenes from the leaves of C. officinalis，and the optimized extraction technology was validated. RESULTS ：The optimal extraction technology of total triterpenes from the leaves of C. officinalis were as follows as ethanol concentration of 73％，liquid-to-material ratio of 38 ∶ 1（mL/g）， extraction time of 60 min. Results of 3 validation tests showed that the contents of total triterpenes from the leaves of C. officinalis were 6.92％，6.91％，6.84％；the average content was 6.89％（RSD＝0.63％），relative error of which with the predicted value （7.28％）was 5.36％. CONCLUSIONS ：The optimized technology is stable and reliable ，and can be used for the extraction of total triterpenes from leaves of C. officinalis .

Optimization of Extraction and Enrichment Process and Anti-hypoxia Activity of Total Triterpenes from Sibiraea Angustata / 中国中医药信息杂志

Objective To optimize the extraction and enrichment process of total triterpenes from Sibiraea Angustata; To study its anti-hypoxia activity. Methods The extraction solvent of total triterpenoids of Sibiraea Angustata was screened by atmospheric pressure anti-hypoxia test. The content of total triterpenes and extraction rate were set as evaluation indexes. The effects of extraction time, extraction times and ratio of material to liquid on extraction were investigated by orthogonal test. The extraction process was optimized. Resin type, concentration of sample, eluent concentration and eluent volume were investigated to optimize the enrichment process of total triterpenes of Sibiraea Angustata. Finally, the anti-hypoxia activity was studied by atmospheric pressure test and acute decompression test. Results The optimum extraction process was as follows: 70% ethanol extraction, the ratio of material to liquid was 1:20, extrat 2 times with 1.5 h for each time, and the content of total triterpenes was 32.25%. The optimum enrichment process was: AB-8-type macroporous adsorption resin, the loading concentration of 2 mg/mL, 80% ethanol 5 BV elution, and total triterpenes content was up to 61.09%; Sibiraea Angustata total triterpenes anti-hypoxia activity test was strong, and had a certain dose dependency. Conclusion The extraction and enrichment process of total triterpenes from Sibiraea Angustata is stable, reproducible and has significant anti-hypoxia activity.

Preparation and dissolusion performance of total triterpenes from Paliurus ramosissimus solid dispersion / 中草药

Objective To prepare total triterpene of Paliurus ramosissimus solid dispersions (TTPR-SD) and study their dissolution properties in vitro. Methods The dissolution of paliurusene and betulinic acid were used as indicators to examine the effect of different carrier materials and different preparation methods on drug dissolution in total triterpenoid solid dispersions by single factor. The preparation technology parameters of total triterpenes of P. ramosissimus solid dispersion were optimized by orthogonal design. Existential state of drugs in this solid dispersion was analyzed by FT-IR and X-ray. Results The solid dispersion was prepared by solvent method with PVPK30 as carrier, the ratio of drug to carrier was 1∶5, ultrasonic for 10 min, and the magnetic stirring temperature was 80 ℃. The cumulative dissolution of paliurusene and betulinic acid within 60 min was achieved 89.19% and 80.49%, respectively. The drug dispersed in the PVPK30 in an amorphous state, and combined with the carrier in the form of hydrogen bonds. Conclusion The prepared solid dispersion can significantly improve the dissolution properties of total triterpene of P. ramosissimus.