Separation,identification,and quantification of active constituents in Fructus Psoraleae by high-performance liquid chromatography with UV,ion trap mass spectrometry,and electrochemical detection

The qualitative and quantitative analysis of active constituents in Fructus Psoraleae is presented by high-performance liquid chromatography (HPLC) coupled with different detections.Extracts of Fructus Psoraleae were examined by HPLC with ion trap mass spectrometry (IT-MS) and 18 major compounds of coumarins,benzofuran glycosides,flavonoids,and meroterpene were identified.The determination of four major constituents including bavachin,isobavachalcone,bavachinin,and bakuchiol was accomplished by HPLC with UV,MS,and electrochemical detection (ECD).These methods were evaluated for a number of validation characteristics (repeatability,LOD,calibration range,and recovery).ECD obtained a high sensitivity for analysis of the four components;MS provided a high selectivity and sensitivity for determination of bavachin,isobavachalcone,and bavachinin in negative-ion mode.After optimization of the methods,separation,identification.and quantification of the four components in Fructus Psoraleae were comprehensively tested by HPLC with UV,MS,and ECD.

Chiral separation of bavachinin in Fructus Psoraleae and rat plasma by liquid chromatography using permethylated-b-CD as a chiral selector

A simple, sensitive and selective method of high-performance liquid chromatography （HPLC） has been successfully developed for separation of bavachinin enantiomers in Fructus Psoraleae and rat plasma. The separation and detection conditions of HPLC were optimized. Chiral bavachinin were separated with the mobile phase of methanol and water （70：30, v/v） at a flow rate of 1.0 mL/min. The linear ranges were in the range of 20-1000 μg/mL. The detection limits were tested as 4 ng/mL and 6 ng/mL for （＋）-bavachinin and （-）-bavachinin, respectively. The method has been applied to analyze chiral bavachinin in rat plasma. HPLC-MS method was used to test the accuracy.

Eucommia and fructus psoraleae promote the proliferation of osteoblasts cultured in vitro: an experimental study

Objective:To study the effect of eucommia and fructus psoraleae treatment on the proliferation of osteoblasts cultured in vitro.Method:1-day-old SD rats were taken, the osteoblasts in exposed bone were separated, cultured and divided into 4 groups, group A were treated with DMEM without drugs or serum, group B were treated with serum-free DMEM containing eucommia with final concentration 10 μg/L, group C were treated with serum-free DMEM containing fructus psoraleae with final concentration 10 μmol/L, and group D were treated with serum-free DMEM containing eucommia with final concentration 10 μg/L and fructus psoraleae with final concentration 10 μmol/L. 24 h after treatment, the mRNA expression of apoptosis genes and proliferation genes in cells were detected.Results: Bax, Fas, FasL and HSG mRNA expression in group B, group C and group D were significantly lower than those in group A while c-fos, c-jun, CyclinD1, Egr-1 and NDRG1 mRNA expression were significantly higher than those in group A;Bax, Fas, FasL and HSG mRNA expression in group D were significantly lower than those in group B and group C, and c-fos, c-jun, CyclinD1, Egr-1 and NDRG1 mRNA expression were significantly higher than those in group B and group C.Conclusion: Both eucommia and fructus psoraleae can promote the osteoblast proliferation, and the combination of the two drugs has synergistic effect.

New incompatible pair of TCM:Epimedii Folium combined with Psoraleae Fructus induces idiosyncratic hepatotoxicity under immunological stress conditions

Epimedii Folium(EF)combined with Psoraleae Fructus(PF)is a common modern preparation,but liver injury caused by Chinese patent medicine preparations containing EF and PF has been frequently reported in recent years.Zhuangguguanjiewan pills(ZGW),which contain EF and PF,could induce immune idiosyncratic liver injury according to clinical case reports and a nonhepatotoxic dose of lipopolysaccharide(LPS)model.This present study evaluated the liver injury induced by EF or PF alone or in combination and investigated the related mechanism by using the LPS model.Liver function indexes and pathological results showed that either EF or PF alone or in combination led to liver injury in normal rats;however,EF or PF alone could lead to liver injury in LPS-treated rats.Moreover,EF combined with PF could induce a greater degree of injury than that caused by EF or PF alone in LPS-treated rats.Furthermore,EF or PF alone or in combination enhanced the LPS-stimulated inflammatory cytokine production,implying that IL-1β,which is processed and released by activating the NLRP3 inflammasome,is a specific indicator of EF-induced immune idiosyncratic hepatotoxicity.Thus,EF may induce liver injury through enhancing the LPS-mediated proinflammatory cytokine production and activating the NLRP3 inflammasome.In addition,the metabolomics analysis results showed that PF affected more metabolites in glycerophospholipid and sphingolipid metabolic pathways compared with EF in LPS model,suggesting that PF increased the responsiveness of the liver to LPS or other inflammatory mediators via modulation of multiple metabolic pathways.Therefore,EF and PF combination indicates traditional Chinese medicine incompatibility,considering that it induces idiosyncratic hepatotoxicity under immunological stress conditions.

Quantitative proteomics analysis of Fructus Psoraleae-induced hepatotoxicity in rats

Fructus Psoraleae,which is commonly consumed for the treatment of osteoporosis,bone fracture,and leucoderma,induces liver injury.This study investigated the pathogenesis of the ethanol extract of Fructus Psoraleae(EEFP)-induced liver injury in rats.EEFP(1.35,1.80,and 2.25 g·kg^–1)was administrated to Sprague Dawley(SD)rats for 30 d.We measured liver chemistries,histopathology,and quantitative isobaric tags for relative and absolute quantitation(iTRAQ)-based protein profiling.EEFP demonstrated parameters suggestive of liver injury with changes in bile secretion,bile flow rate,and liver histopathology.iTRAQ analysis showed that a total of 4042 proteins were expressed in liver tissues of EEFP-treated and untreated rats.Among these proteins,81 were upregulated and 32 were downregulated in the treatment group.KEGG pathway analysis showed that the drug metabolic pathways of cytochrome P450,glutathione metabolism,glycerolipid metabolism,and bile secretion were enriched with differentially expressed proteins.The expression of key proteins related to the farnesoid X receptor(FXR),i.e.,the peroxisome proliferators-activated receptor alpha(PPAR-α),were downregulated,and multidrug resistance-associated protein 3(MRP3)was upregulated in the EEFP-treated rats.Our results provide evidence that EEFP may induce hepatotoxicity through various pathways.Furthermore,our study demonstrates changes in protein regulation using iTRAQ quantitative proteomics analysis.

Landscape of Hepatobiliary Adverse Drug Reactions Related to Preparations Containing Psoraleae Fructus and Its Application in Pharmacovigilance

Objective To analyze clinical feature and information of medication to explore the risk signals of preparations containing Psoraleae Fructus(BGZP)related with hepatobiliary adverse drug reactions(ADR),in order to reinforce pharmacovigilance.Methods A retrospective study was conducted based on hepatobiliary ADR related with BGZP from the China Adverse Drug Reaction Monitoring System in years from January 2012 to December 2016.Serious and general ADRs were analyzed and assessed.Results There were 355 cases of hepatobiliary ADR related to BGZP.Both the amount of cases and the proportion of serious ADR showed an increasing growth by years(P<0.05).It was found that 10.43%of 355 cases may be involved with irrational drug use,including overdose,repeated medication,and combination of multiple drugs.There were 190 cases which used BGZP(non-combination),and they were mainly for common in diseases caused by abnormal immune activation(accounting for 40.53%of the total cases).Especially at the age group with the most cases with age of 41–50 years,the cases associated with immunological diseases of female were obviously more than that of male(P<0.05).The latency of hepatobiliary ADR related to BGZP ranged from 1 to 386 days,and the median latency was 27.5 days,along with the range of cumulative dose(0.45–520.02 g)as well as the daily dose(0.09–2.64 g/d)after the conversion.Conclusions Cases of hepatobiliary ADR related to BGZP showed significant individual differences,and there was no correlation between drug usage duration and dosage and the occurrence of hepatobiliary ADR.It may be similar with idiosyncratic drug-induced liver injury,and recommended that BGZP should be used with more caution under monitoring liver function,especially in female patients with immunological diseases.

Interactions of drug-metabolizing enzymes with the Chinese herb Psoraleae Fructus

Psoraleae Fructus(the dried fruits of Psoralea corylifolia), one of the most frequently used Chinese herbs in Asian countries, has a variety of biological activities. In clinical settings, Psoraleae Fructus or Psoraleae Fructus-related herbal medicines frequently have been used in combination with a number of therapeutic drugs for the treatment of various human diseases, such as leukoderma, rheumatism and dysentery. The use of Psoraleae Fructus in combination with drugs has aroused concern of the potential risks of herb-drug interactions(HDI) or herb-endobiotic interactions(HEI). This article reviews the interactions between human drug-metabolizing enzymes and the constituents of Psoraleae Fructus;the major constituents in Psoraleae Fructus, along with their chemical structures and metabolic pathways are summarized, and the inhibitory and inductive effects of the constituents in Psoraleae Fructus on human drug-metabolizing enzymes(DMEs), including target enzyme(s), its modulatory potency, and mechanisms of action are presented. Collectively, this review summarizes current knowledge of the interactions between the Chinese herb Psoraleae Fructus and therapeutic drugs in an effort to facilitate its rational use in clinical settings, and especially to avoid the potential risks of HDI or HEI through human DMEs.

Pancreatic lipase inhibitory constituents from Fructus Psoraleae

Pancreatic lipase(PL), a crucial enzyme in the digestive system of mammals, has been proven as a therapeutic target to prevent and treat obesity. The purpose of this study is to evaluate and characterize the PL inhibition activities of the major constituents from Fructus Psoraleae(FP), one of the most frequently used Chinese herbs with lipid-lowering activity. To this end, a total of eleven major constituents isolated from Fructus Psoraleae have been obtained and their inhibition potentials against PL have been assayed by a fluorescence-based assay. Among all tested compounds, isobavachalcone, bavachalcone and corylifol A displayed strong inhibition on PL(IC50 < 10 μmol·L-1). Inhibition kinetic analyses demonstrated that isobavachalcone, bavachalcone and corylifol A acted as mixed inhibitors against PL-mediated 4-methylumbelliferyl oleate(4-MUO) hydrolysis, with the Ki values of 1.61, 3.77 and 10.16μmol·L-1, respectively. Furthermore, docking simulations indicated that two chalcones(isobavachalcone and bavachalcone) could interact with the key residues located in the catalytic cavity of PL via hydrogen binding and hydrophobic interactions. Collectively, these finding provided solid evidence to support that Fructus Psoraleae contained bioactive compounds with lipid-lowering effects via targeting PL, and also suggested that the chalcones in Fructus Psoraleae could be used as ideal leading compounds to develop novel PL inhibitors.

A systematic review on the safety of Psoraleae Fructus:potential risks,toxic characteristics,underlying mechanisms and detoxification methods

Psoraleae Fructus(PF)is an important traditional herbal medicine with a long history of clinical application.It is widely used to treat various diseases,such as osteoporosis,leucoderma and diarrhea.As a traditional nontoxic herb,it has aroused worldwide concern about the potential risks due to increasing adverse reaction events.This article reviews the botany,ancient records of medical uses,adverse reactions,toxicological research advance and detoxification methods of PF.According to clinical studies,liver injury is the most predominant in PF-related adverse reactions.The underlying mechanisms include bile acid metabolism and transport disorders,oxidative stress,mitochondrial damage,inhibition of liver cell regeneration and inflammatory reactions.Furthermore,the potential toxins of PF are summarized.Traditional methods of processing and compatibility will provide reference for reducing the toxicity of PF,which requires further research.In sum,this work systematically summarizes the reserach progress on the safety of PF,which will provide comprehensive insights into the toxicity of PF and facilitate its safe use and future development.

补骨脂种子提取物对茶饼病菌的抑菌活性

探讨补骨脂(Psoralea corylifolia L.)种子提取物对茶饼病菌(Exobasidium vexans Massee)的抑菌活性及田间防治效果。采用孢子萌发抑制率法和菌丝生长速率法测定补骨脂种子提取物的抑菌活性,按照农药田间药效试验准则在5省5地进行茶饼病田间试验。室内抑菌活性试验结果表明,补骨脂种子提取物对茶饼病菌孢子萌发和菌丝生长具有明显的抑制作用,抑制率随着药剂浓度的增加而增加,其EC50分别为12.925 mg/L和6.074 mg/L。田间试验结果表明,当补骨脂种子提取物微乳剂有效成分用量为2.00~2.67 mg/kg时,其对茶饼病的防治效果为68.08%~83.78%,其防治效果相当或优于对照药剂3%多抗霉素可湿性粉剂。

Quality Evaluation and Regional Analysis of Psoraleae Fructus by HPLC-DAD-MS/MS plus Chemometrics

Objective To evaluate the quality of Psoralea corylifolia collected from 12 provinces of China. Methods An HPLC-DAD-MS/MS method was used to identify, determine, and estimate 14 representative bioactive compounds in P. corylifolia. Then on the basis of the content data, the chemometrics method was used to differentiate 20 samples from different regions. Results The quality of P. corylifolia from 12 different provinces of China was evaluated by this method. Though the samples showed similar profiles, content of the detected markers varied significantly in different regions and batches. According to the results of the hierarchical cluster analysis and principle component analysis, it can be concluded that the samples from different origins could be clustered reasonably into two groups, as well as successfully distinguished. Conclusion A simple and reliable new method which used HPLC-DAD-MS/MS and chemometrics has been developed to characterize, classify, and control the quality of P. corylifolia.

补骨脂盐炙前后HPLC指纹图谱的比较

建立补骨脂(Psoralea corylifolia L.)盐炙前后的高效液相(HPLC)指纹图谱,并采用化学计量学方法评价生品补骨脂、盐补骨脂的差异。采用HPLC法建立指纹图谱,色谱柱为Waters Symmetry C18柱(4.6 mm×150 mm,5μm),流速为1.0 mL/min,柱温为30℃,进样量为10μL,检测波长为290 nm。采用镜像对比、偏最小二乘判别(PLS-DA)、相似性评价分析方法进行数据分析。成功建立了生品补骨脂、盐补骨脂的HPLC指纹图谱,共确定14个共有峰,并指认了补骨脂素、异补骨脂素2种成分。生品补骨脂相似度良好,盐炙后的补骨脂相似度发生变化。镜像分析结果表明,盐炙后的补骨脂HPLC指纹图谱与生品补骨脂存在差异。与生品补骨脂相比,盐补骨脂的1~14号峰均有不同程度的增加。PLS-DA分析结果表明,生品补骨脂与盐补骨脂在化学成分上存在差异。建立的指纹图谱方法重复性、稳定性良好,可为补骨脂盐炙前后的鉴别及质量分析提供参考。

EFFECTS OF BU GU ZHI(PSORALEA CORYLIFOLIA L) AND BAI ZHI(DAHURIAN ANGELICA ROOT) EXTRACTS ON MELANOCYTE ADHESION AND MIGRATION

Objective To find direct effect of Chinese herbs Bu Gu Zhi (Psoralea corylifolia L) and Bai Zhi (Dahurian angelica root) Extracts on melanocyte adhesion and migration in vitro. Methods Ethanol extracts obtained from two kind of Chinese medicinable herbs were tested. Human melanocytes were obtained from neonatal foreskins and 48-well culture dish covered with fibronectin were used to perform melanocyte adhesion assay; Motility was assessed using the micropore filter method. Results: The extracts of Bu Gu Zhi(Psoralea corylifolia L), Bai Zhi(Dahurian angelica root) obviously showed an effect in increasing of human melanocyte adhesion and migration on fibronectin. Conclusion It is suggested that Buguzhi(Fructus Psoraleae) and Baizhi(Radix Angelicae Dahuricae) might induce melanocyte adhesion and/or migration in the treatment of vitiligo.

Pharmacokinetic study about compatibility of Eucommia ulmoides and Psoralea corylifolia

Objective:The compatibility of Eucommia ulmoides(Eu)and Psoralea corylifolia(Pc)on the pharmacokinetic(PK)properties in the rat was explored in this study.Methods:Eu extract,Pc extract and the combined extracts(crude drug ratio was 2:1)was administered by gavage,respectively.Two PK experiments were conducted.In first one,the blood samples were collected via the occuli chorioideae vein to get the PK properties of the components.In second one,the blood samples were simultaneously collected via the internal jugular vein or portal vein at different time points and the concentrations of target ingredients were detected by LC/MS/MS to clear the location where the interaction of Eu and Pc took place in vivo.Results:Eight of 11 ingredients in Eu and Pc extract were determined in rat plasma.The exposure levels of geniposidic acid(GPA),aucubin(AU),geniposide(GP),pinoresinol diglucoside(PDG),psoralen glycosides(PLG)and isopsoralen glycosides(IPLG)were decreased 1/5–2/3 after administration of combined extracts.Comparing to the combined administration,the exposure of GPA and AU in plasma of single Eu administration collected via the portal vein were decreased 1/3–2/3,and the values of AUC0-24h and AUC0-∞ of GP collected from the portal vein or internal jugular vein were double increased.The other components’parameters were not significantly changed.Conclusion:In summary,the Pc and Eu combined administration could affect the exposure of the main components of Eu extract in rats due to the changed intestinal absorption.The research on the compatibility of Pc and Eu was helpful to guide the clinical administration of Eu and Pc simultaneously.

Bavachin enhances NLRP3 inflammasome activation induced by ATP or nigericin and causes idiosyncratic hepatotoxicity

Psoraleae Fructus(PF)is a well-known traditional herbal medicine in China,and it is widely used for osteoporosis,vitiligo,and other diseases in clinical settings.However,liver injury caused by PF and its preparations has been frequently reported in recent years.Our previous studies have demonstrated that PF could cause idiosyncratic drug-induced liver injury(IDILI),but the mechanism underlying its hepatotoxicity remains unclear.This paper reports that bavachin isolated from PF enhances the specific stimuli-induced activation of the NLRP3 inflammasome and leads to hepatotoxicity.Bavachin boosts the secretion of IL-ip and caspase-1 caused by ATP or nigericin but not those induced by poly(I:C),monosodium urate crystal,or intracellular lipopolysaccharide.Bavachin does not affect AIM2 or NLRC4 inflammasome activation.Mechanistically,bavachin specifically increases the production of nigericin-induced mitochondrial reactive oxygen species among the most important upstream events in the activation of the NLRP3 inflammasome.Bavachin increases the levels of aspartate transaminase and alanine aminotransferase in serum and hepatocyte injury accompanied by the secretion of IL-ip via a mouse model of lipopolysaccharide-mediated susceptibility to IDILI.These results suggest that bavachin specifically enhances the ATP-or nigericin-induced activation of the NLRP3 inflammasome.Bavachin also potentially contributes to PF-induced idiosyncratic hepatotoxicity.Moreover,bavachin and PF should be evaded among patients with diseases linked to the ATP-or nigericin-mediated activation of the NLRP3 inflammasome,which may be a dangerous factor for liver injury.