Therapeutic effect of Leonuri Herba aqueous decoction on primary dysmenorrhea in rats and its metabolomic analysis / 中国中药杂志

This study aimed to investigate the therapeutic effect of Leonuri Herba aqueous decoction on primary dysmenorrhea(PD) and explore the underlying mechanism in conjunction with untargeted metabolomics. Forty adult female rats were randomly divi-ded into a normal group, a model control group, ibuprofen(0.12 g·kg~(-1)) group, and high-and low-dose Leonuri Herba aqueous decoction(5 and 2.5 g·kg~(-1)) groups, with eight rats in each group. The PD rat model was prepared using intramuscular injection of estradiol benzoate combined with intraperitoneal injection of pitocin. Drugs were administered by gavage from the 4th day of modeling for 7 d. After the last administration, pitocin was injected intraperitoneally, and the writhing latency and writhing times within 30 min were recorded. The uterine and ovarian coefficients were determined. Estradiol(E_2), progesterone(Prog), oxytocin(OT), cyclooxyge-nase 2(COX-2), prostaglandin E_2(PGE_2), prostaglandin F_(2α)(PGF_(2α)), and Ca~(2+) levels in uterine tissues were measured by ELISA and biochemical kits. Morphological changes in uterine and ovarian tissues were observed by hematoxylin-eosin(HE) staining. The protein expression of oxytocin receptor(OTR), prostaglandin E_2 receptor 3(EP3), and estrogen receptor alpha(ERα) in uterine tissues was detected by immunohistochemistry. The mRNA expression of OTR, PGE_2 receptors 1-4(EP1, EP2, EP3, and EP4), and PGF_(2α) receptor(FP) in uterine tissues was detected by quantitative real-time PCR. Untargeted metabolomics analysis was performed by ultra-high-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry(LC-QTOF-MS) technology to screen potential biomarkers and enrich metabolic pathways. The results showed that Leonuri Herba was able to significantly reduce the writhing times in PD rats(P<0.05 or P<0.01), significantly reduce the uterine and ovarian coefficients(P<0.01), and improve their histomorphology. After treatment with Leonuri Herba, PGE_2 content was significantly increased(P<0.05), COX-2, PGF_(2α) and Ca~(2+) content, and PGF_(2α)/PGE_2 was significantly decreased(P<0.05 or P<0.01), and OT content was decreased, while E_2 and Prog content tended to further increase in uterine tissues of PD rats. Correspondingly, OTR and EP3 protein expression was significantly downregulated(P<0.05 or P<0.01) and ERα protein expression was upregulated(P<0.05) in uterine tissues. The mRNA expression of FP and EP4 in uterine tissues was significantly downregulated(P<0.01), and the mRNA expression of EP1, EP3, and OTR showed a decreasing trend. The untargeted metabolomics results showed that 10 differential metabolites were restored in the plasma of PD rats after Leonuri Herba treatment. The results indicate that Leonuri Herba is effective in the prevention and treatment of PD, and the underlying mechanism may be attributed to the regulation of PGs synthesis and corresponding receptor binding.

Comparison Between Traditional and Integrated Processing Technology of Leonuri Herba Based on Chemical Composition and Pharmacological Effect / 中国实验方剂学杂志

Objective: To elucidate the rationality of integrated processing technology of Leonuri Herba based on comparison of chemical constituents and pharmacological effect of Leonuri Herba between traditional and integrated processing technology. Method:The contents of stachydrine hydrochloride,leonurine hydrochloride,rutin,hyperoside and isoquercetin were used as indexes to compare the differences in the contents of chemical constituents between traditional and integration processing technology of Leonuri Herba.Effect of Leonuri Herba with different processing technology on auricular swelling induced by dimethylbenzene in mice were observed to compared the differences of their anti-inflammatory effect.And rat acute blood stasis model was used to compare the differences of Leonuri Herba with different processing technology on hemorheology and blood coagulation indexes. Result:Contents of stachydrine hydrochloride,leonurine hydrochloride,rutin,hyperoside and isoquercetin in products of integrated processing were 1.558%,0.168%,0.137%,0.113% and 0.078%,they were 1.482%,0.134%,0.125%,0.082% and 0.071% in products of traditional processing,respectively.Both of the processing methods could reduce the degree of swelling and the contents of tumor necrosis factor(TNF)-α,interleukin(IL)-1β,IL-6 in mouse serum.And both of the processing methods could reduce the whole blood viscosity,prolong the thrombin time(TT),prothrombin time(PT),activated partial thromboplastin time(APTT) and reduce the concentration of plasma fibrinogen(FIB) in acute blood stasis model rats. Conclusion:Compared with the traditional processing technology,the integrated processing technology is better in guaranteeing the quality of Leonuri Herba decoction pieces and reducing the production cost,which indicates that the integrated processing of Leonuri Herba is reasonable.

Standard decoction of Leonuri Herba / 中国中药杂志

To build an evaluation standard for quality of Leonuri Herba standard decoction. 13 batches of Leonuri Herba standard decoction with different quality were prepared. The contents of leonurine hydrochloride and stachydrine hydrochloride were determined; then the transfer rate and the extract rate were calculated and pH value was measured; and HPLC fingerprint method was established for analysis. The results of 13 batches of samples revealed that the transfer rate of leonurine hydrochloride and stachydrine hydrochloride was 30.0%-53.4% and 67.0%-82.6%, respectively; the extract rate was 12.1%-18.3% and the pH value was 5.87-6.22. Moreover, 12 common chromatographic peaks were determined based on fingerprint by using Similarity Evaluation System for Chromatographic fingerprint of Traditional Chinese Medicine (2012A). The similarity of 13 batches of samples was analyzed and compared, and the results showed that the similarity was higher than 0.9. In this study, the preparation method for Leonuri Herba decoction was standard, with high similarity in fingerprint, showing high precision, stability and repeatability in fingerprint analysis. Thus, this study can provide a reference for the quality control of Leonuri Herba dispensing granules.

Determination of stachydrine hydrochloride and leonurine hydrochloride in Yimucao preparations by HPLC-MS / 中国中药杂志

To establish the quantitative method of stachydrine hydrochloride and leonurine hydrochloride in the preparations of Leonuri Herba. The contents of stachydrine hydrochloride and leonurine hydrochloride in the preparations of Leonuri Herba were determined by HPLC-MS. The chromatographic column was Waters XBridge Amide(4.6 mm×250 mm,5 μm). The mobile phase was acetonitrile-0.1% formic acid in gradient mode,at the flow rate of 1.0 mL• min⁻¹,with the split ratio of 1∶4. MS conditions for the ESI ion source,positive ion mode,selective ion scan(SIM) of stachydrine hydrochloride(m/z 144.0) and leonurine hydrochloride(m/z 312.0) was measured. The linear ranges of stachydrine hydrochloride was 0.562 8-281.4 μg•L-1（r=0.999 8）. The linear ranges of leonurine hydrochloride was 0.521 2-260.6 μg•L-1（r=0.999 8）. The method is accurate,simple,and reliable,and can be used to determine the contents of stachydrine hydrochloride and leonurine hydrochloride in the preparations of Leonuri Herba.

Effect of Leonuri Herba alkaloids on prostatic hyperplasia in older rats / 中草药

Objective: To investigate the pharmacological effects and mechanism of Leonuri Herba alkaloids (LHA) on prostate hyperplasia in older rats. Methods: The prostate hyperplasia model in older rats was induced by male and female hormone, the rats were divided into five groups: 50.0, 25.0, 12.5 mg/kg alkaloids dosage group, 30 mg/kg Longbishu Capsule suspension group, and model group given the same volume of saline. Otherwise aged rats and young rats were set as control groups administered with saline, and administered once daily for consecutive 30 d. The wet weight and prostate index in rat prostate were measured, and the expression of E2, DHT, T, bFGF, TGF-β1, EGF, and IGF-1 in serum and prostate tissue were detected after the experiment. The rat prostate tissue morphology changes and rat prostate cells ultrastructural changes were observed by light microscopy and transmission electron microscopy. Results: Compared with the model group, benign prostatic hyperplasia rat model was replicated successfully, low-dose LHA can significantly reduce the rat prostate wet weight and prostate index (P < 0.05), mid-dose LHA can significantly reduce the rat prostate index (P < 0.05). Each dose of LHA can significantly reduce the levels of T and DHT in prostate model rats (P < 0.01). Each dose of LHA can significantly reduce the expression of bFGF, EGF, and IGF-1 in prostate model rats (P < 0.01). Low-dose LHA can significantly increase the expression of TGF-β1 in prostate tissue. Each dose of LHA can significantly reduce the density of prostate model rats (P < 0.05, 0.01), and significantly increase the membrane surface and surface area values (P < 0.01). Each dose of LHA can significantly reduce intracytoplasmic mitochondria in prostate cells caused by model, and relieve the pathological changes of mitochondria cristae. Conclusion: LHA has good therapeutic effect to prostatic hyperplasia model rats induced by male and female hormone.