Interaction study of salvianolic acids for injection on pharmacokinetics of clopidogrel in rats using LC-MS/MS.

Salvianolic acids for injection (SAI) is developed from traditional Chinese medicine and approved for the treatment of cardiovascular and cerebrovascular diseases. Clopidogrel is an inhibitor of platelet aggregation, which is often prescribed for patients in combination with SAI. This present study aimed to assess the effects of SAI on the pharmacogenomics, pharmacokinetics, and pharmacodynamics of clopidogrel, thereby ensuring the safety and efficacy of coadministration. In vitro cytochrome P450 isoenzyme assays were performed in human liver microsomes using LC-MS/MS method to assess the metabolites of CYPs substrates. The effects of SAI on the pharmacokinetic and pharmacodynamic behaviors of clopidogrel were investigated in rats. The main pharmacokinetic parameters were analyzed using LC-MS/MS. Prothrombin time, activated partial thromboplastin time, bleeding time, and inhibition of platelet aggregation were measured to evaluate the effects of pharmacodynamics. Our study revealed that the clinical dose of SAI has no significant inhibitory effect on clopidogrel-related liver microsome metabolic CYP450 isoenzymes. Moreover, SAI did not affect the pharmacokinetics of clopidogrel when rats were administered both single and multiple doses. In pharmacodynamic study, SAI has no effect on platelet aggregation rate, prothrombin time, and activated partial thromboplastin time of clopidogrel but could significantly prevent the risk of bleeding caused by clopidogrel.

Ginsenoside Rb1 alleviates liver injury induced by 3-chloro-1,2-propanediol by stimulating autophagic flux.

In recent years, foodborne pollutants have become a hot issue in the field of food safety. 3-chloro-1,2-propanediol (3-MCPD) is a widely existing food contaminant. In our previous study, it was confirmed that 3-MCPD can block autophagic flux by inhibiting lysosomal function, thus causing liver injury. Ginseng is a traditional Chinese herbal medicine that contains a variety of bioactive ingredients, among which ginsenoside Rb1 (Gs-Rb1) is the most abundant. In this study, we aim to use Gs-Rb1 to improve 3-MCPD-induced autophagic flux blockage to alleviate liver injury. First, a nontoxic dose of Gs-Rb1 was identified by screening with the MTT method in which Gs-Rb1was added to HepG2 cells and co-treated with 3-MCPD. We found that Gs-Rb1 effectively enhanced the cell activity inhibited by 3-MCPD. Meanwhile, apoptosis data showed that Gs-Rb1 significantly alleviated the apoptosis of HepG2 cells induced by 3-MCPD. Subsequently, we found that Gs-Rb1 could alleviate autophagic flux blockage caused by 3-MCPD in a dose-dependent manner by detecting autophagy-related protein levels and transfecting mRFP-GFP-LC3 adenovirus. On this basis, we used Western blotting and qPCR to explore whether miR-128 was involved in the alleviation effect of Gs-Rb1 on autophagic flux blockade induced by 3-MCPD. The results showed that Gs-Rb1 inhibited the expression of miR-128 and promoted the nuclear expression and target gene transcription of TFEB. Finally, the findings were confirmed by using a hsa-miR-128 inhibitor and mimic. We found that hsa-miR-128 inhibitor alleviated the autophagic flux blockage and apoptosis caused by 3-MCPD and Gs-Rb1 also had a certain alleviation effect on the autophagic flux blockage and apoptosis caused by hsa-miR-128 mimic. This study elaborated the mechanism by which Gs-Rb1 alleviates hepatotoxicity induced by foodborne 3-MCPD by stimulating autophagic flux via miR-128-targeted TFEB, which provides a reliable theoretical basis and target for the use of natural substances to reduce the harm of food processing pollutants on the human body. PRACTICAL APPLICATION: We found that natural ginsenoside Rb1 can alleviate liver injury induced by 3-MCPD(a toxic substance found in foods such as refined vegetable oil, soy sauce, and baby milk powder), which is conducive to the development and utilization of ginseng and has practical significance for the prevention of foodborne liver injury.

Pharmacokinetics, tissue distribution and excretion of a novel long-acting human insulin analogue - recombinant insulin LysArg in rats.

As a novel long-acting recombinant human insulin analogue, it is necessary to carry out the preclinical research for insulin LysArg. The purpose of this study was to characterise the pharmacokinetic, tissue distribution and excretion of insulin LysArg and provide a reference for its development. Three methods were used to measure the content of insulin LysArg in biological samples after a single subcutaneous administration in rats, including radioassay, radioassay after precipitation with TCA and separation by HPLC. After Subcutaneous administration of recombinant insulin LysArg 1, 2, 4 U/kg in rats, it showed both Cmax and AUC0-t were positively correlated with the dose. In the meanwhile, after a single subcutaneous administration of recombinant insulin LysArg at 2 U/kg in rats, the amount of radioactivity in most organs was highest at 1.5 h and then decreased gradually, no accumulation was found. The highest level of insulin LysArg was observed in the kidney. Like other macromolecules, insulin LysArg was mainly excreted from urine. The study fully illustrated the pharmacokinetic pattern of insulin LysArg, provided valuable informations to support its further development about safety and toxicology.

Transporters (OATs and OATPs) contribute to illustrate the mechanism of medicinal compatibility of ingredients with different properties in yuanhuzhitong prescription.

Various medicinal ingredients with different tastes are combined according to the theory of compatibility in Chinese materia medica to achieve a better efficacy, while the mechanism was not very clear. Here, the authors studied the interaction between ingredients and human transporters such as the kidney transporters OAT1 and OAT3, the liver transporters OATP1B1 and OATP1B3, and the intestine transporter OATP2B1 to discern the compatibility mechanism of ingredients with different tastes in the Yuanhuzhitong preparation (YHP) comprising Corydalis yanhusuo (CYH) and Angelica dahurica (AD), which could relieve pain by restraining the central system. The results show that tetrahydropalmatine (TDE), the major component of CYH, could be transported by OAT3 into kidney, OATP1B1 and OATP1B3 into liver, while imperatorin (IPT) and isoimperatorin (ISP), the two key components of AD, and AD extract showed strong inhibition to OAT1 and OAT3. What's more, AD extract also exerted strongly inhibition to human transporters OATP1B1 and OATP1B3. It was also detected that IPT, ISP, and AD extract significantly downregulated the expression of Oatp1a1, Oatp1a4, and Oatp1b2 of liver in mice. The in vivo results show that the concentration of TDE in liver and kidney significantly decreased, while the TDE concentration in blood and brain were both significantly enhanced in the presence of IPT, ISP, and AD extract. These results suggest that the ingredients in AD with pungent taste could enhance the exposure of TDE in blood and brain by inhibiting the uptake of TDE in liver and kidney. That is to say, TDE with bitter taste could "flood up" into the central nervous system to play its therapeutic effect by the cut-off of that into liver and kidney in the presence of ingredients within AD. This paper not only proves the meridian distribution of CYH in liver and kidney with the role of OAT3, OATP1B1, and OATP1B3, but also illustrates how to improve the efficacy of CYH by reasonable compatibility with AD. This study may offer a valuable clue to illustrate the mechanism of compatibility theory.

Nobiletin potentiates paclitaxel anticancer efficacy in A549/T xenograft model: Pharmacokinetic and pharmacological study.

BACKGROUND: Nobiletin (N), a polymethoxylated flavone from citrus fruits, enhanced anti-cancer effects of paclitaxel (PTX) in multi-drug resistance (MDR) cancer cells via inhibiting P-glycoprotein (P-gp) in our previous report. But the in vivo chemo-sensitizing effect of nobiletin is unknown. Moreover, considering the nonlinear pharmacokinetics and narrow therapeutic window of PTX, drug-drug interaction should be explored for using nobiletin with PTX together. PURPOSE: In this study, we wanted to explore whether nobiletin could affect the pharmacokinetic (PK) behavior of PTX and reverse drug resistance in vivo as well as the corresponding mechanisms. STUDY DESIGN AND METHODS: Accurate and sensitive UPLC-MS/MS method was developed for the detection of PTX, and was applied to the pharmacokinetic study in rats. In vivo anti-MDR tumor study was carried out with A549/T xenograft nude mice model. Immunohistochemistry and western blot analysis were used for evaluating the levels of P-gp, Nrf2, and AKT/ERK pathways in MDR tumors. RESULTS: Nobiletin significantly enhanced the therapeutic effects of PTX, and inhibited the MDR tumor sizes in the A549/T xenograft model, while PTX or nobiletin alone did not. We found that nobiletin increased the PTX concentrations in tumor tissues but did not affect the PK behavior of PTX. Notably, Nrf2 and phosphorylation of AKT/ERK expression in MDR tumor tissues were significantly inhibited by giving nobiletin and PTX together. However, nobiletin did not affect the expression of P-gp. CONCLUSION: Nobiletin reversed PTX resistance in MDR tumor via increasing the PTX content in the MDR tumor and inhibiting AKT/ERK/Nrf2 pathways, but without affecting the systematic exposure of PTX, indicating that nobiletin may be an effective and safe MDR tumor reversal agent.

Effects of liquorice on pharmacokinetics of aconitine in rats.

Aconite alkaloids are the main bioactive ingredients existing in Aconitum, for instance aconitine (AC), which exhibit potent analgesic, antirheumatic and other pharmacological effects. In this study, effects of long-term treatment with liquorice on pharmacokinetics of AC in rats were investigated. Pharmacokinetics of AC after oral administration of AC at 1.5 mg/kg either with pre-treatment of liquorice water extracts at 0.433 or 1.299 g/kg (crude drug), respectively, for one week or not were studied. Additionally, LS-180 cells and human primary hepatocytes were utilized to explore the potential effects of bioactive ingredients of liquorice on P-glycoprotein (P-gp) and Cytochromes P450 (CYPs), respectively. The results revealed that exposure of AC after pre-treatment with liquorice was altered remarkably. Area under the concentration-time curve (AUC) decreased from 161 ± 37.8 to 58.8 ± 8.97 and 44.7 ± 8.20 ng/mL*h, respectively. Similarly, Cmax decreased from 26.2 ± 5.19 to 11.8 ± 1.15 and 6.86 ± 0.600 ng/mL, respectively. In addition, expressions of CYPs of human primary hepatocytes were enhanced to various contents after induction. Moreover, accumulation of AC and hypaconitine (HA), not mesaconitine (MA) inside of LS-180 cells were reduced after pre-treatment by comparison with control. In conclusion, the exposure of AC in vivo declined after pre-treatment with liquorice extract, which may be highly associated with upregulated expression and/or function of CYPs and P-gp.

Potential detoxification effect of active ingredients in liquorice by upregulating efflux transporter.

BACKGROUND: As one of most widely used herbal medicine, liquorice exhibits diverse pharmacological activities, for instance, analgesic, antitussive, antiarrhythmic, anti-inflammatory, and immune regulation. Additionally, detoxification effects were observed in combination of liquorice with other herbal drugs. The mechanism of detoxification of liquorice has been extensively investigated through material basis and interference with CYPs though, investigations of its effect on transporters were very limited, according to the literature. PURPOSE: The objective of this study was attempt to investigate the effect of active ingredients existing in liquorice on the efflux transporters as to clarify the potential mechanism of detoxification of liquorice. METHODS: Multiple analytical approaches have been explored, including flow cytometry, fluorescent detection, RT-PCR, Western blot to measure the function, activity as well as mRNA/protein expression of efflux transporters on LS-180 cell model after treatments with active compounds of liquorice. Additionally, Caco-2 cell model was utilized to further investigate the potential impact of those ingredients on efflux transporter. RESULTS: The resulting data indicated that those active ingredients, including flavonoids (liquiritin, liquiritigenin, isoliquiritin, isoliquiritigenin and licochalcone A) and pentacyclic triterpene saponin (glycyrrhetinic acid) were able to upregulate the expression of efflux transporters, for example P-gp, BCRP and MRP2. The gene expressions were approximately over 2.5 folds by comparison with that of control, and up to 13 folds and 16 folds for BCRP by isoliquiritin and isoliquiritigenin, and further confirmed by Western blot. The functional assay also supported up-regulation of efflux transporter by those ingredients. Flow cytometry study showed that the level of rhodamine123 as probe substrate in LS-180 cells decreased to approximately 50% after treatment with active ingredients of liquorice, compared with that of control. The fluorescent assay confirmed that change of rhodamine 123 was correlated with the concentrations of active ingredients given. The efflux transport of rhodamine 123 was enhanced in Caco-2 cell models as well. CONCLUSION: The study clarified potential detoxification mechanism of liquorice by up-regulating efflux transporter as to reduce absorption of xenobiotics across small intestinal membrane, which provided a new insight into pharmacological function of liquorice.

An integrated approach to uncover quality marker underlying the effects of Alisma orientale on lipid metabolism, using chemical analysis and network pharmacology.

BACKGROUND: Quality control of traditional Chinese medicines is currently a great concern, due to the correlation between the quality control indicators and clinic effect is often questionable. According to the "multi-components and multi-targets" property of TCMs, a new special quality and bioactivity evaluation system is urgently needed. PURPOSE: Present study adopted an integrated approach to provide new insights relating to uncover quality marker underlying the effects of Alisma orientale (AO) on lipid metabolism. METHODS: In this paper, guided by the concept of the quality marker (Q-marker), an integrated strategies "effect-compound-target-fingerprint" was established to discovery and screen the potential quality marker of AO based on network pharmacology and chemical analysis. Firstly, a bioactivity evaluation was performed to screen the main active fractions. Then the chemical compositions were rapidly identified by chemical analysis. Next, networks were constructed to illuminate the interactions between these component and their targets for lipid metabolism, and the potential Q-marker of AO was initially screened. Finally, the activity of the Q-markers was validated in vitro. RESULTS: 50% ethanol extract fraction was found to have the strongest lipid-lowering activity. Then, the network pharmacology was used to clarify the unique relationship between the Q-markers and their integral pharmacological action. CONCLUSION: Combined with the results obtained, five active ingredients in the 50% ethanol extract fraction were given special considerations to be representative Q-markers: Alisol A, Alisol B, Alisol A 23-acetate, Alisol B 23-acetate and Alisol A 24-acetate, respectively. The chromatographic fingerprints based Q-marker was establishment. The integrated Q-marker screen may offer an alternative quality assessment of herbal medicines.

Identify super quality markers from prototype-based pharmacokinetic markers of Tangzhiqing tablet (TZQ) based on in vitro dissolution/ permeation and in vivo absorption correlations.

BACKGROUND: A quality marker (Q-marker) is defined as an inherent chemical compound that is used for the quality control of a drug. Its biological activities are closely related to safety and therapeutic effects. Generally, a multiple-component herbal medicine may have many Q-markers. We therefore proposed a concept of "super Q-marker" satisfying both the criterion of Q-markers and PK-markers to be used in more effective quality control of herbal medicine. PURPOSE: The first aim was to find suitable prototype-based PK-markers from Tangzhiqing tablets (TZQ), a Chinese patent medicine. Then super Q-markers were expected to be identified from the prototype-based PK-markers based on an in vitro-in vivo correlation study. METHODS: Potentially eligible prototype-based PK-markers were identified in a single- and multiple-dose pharmacokinetic study on TZQ in 30 healthy volunteers. The in vitro dissolution and permeation profiles of the prototype-based PK-markers of TZQ were evaluated by the physiologically-based drug dissolution/absorption simulating system (DDASS). An in vitro-in vivo correlation analysis was conducted between the dissolution/permeation behaviors in DDASS and the actual absorption profiles in human to test the transferability and traceability of the promising super Q-markers for TZQ. RESULTS: In human, plasma paeoniflorin and nuciferine as prototype-based PK-markers exhibited the appropriate pharmacokinetic properties, including dose-dependent systemic exposure (AUC, Cmax) and a proper elimination half-life (1∼3h). In DDASS, it was predicted that paeoniflorin and nuciferine are highly permeable but the absorption rates are primarily limited by the dissolution rates. Moreover, the established in vitro-in vivo correlations of paeoniflorin and nuciferine were in support of the super Q-markers features. CONCLUSION: Paeoniflorin and nuciferine are identified as the super Q-markers from the prototype-based PK-markers of TZQ based on findings from a combination of in vitro, in vivo, and in vitro-in vivo correlation studies. This method is practical for optimal identification of qualified Q-markers, thus helping improve the quality control of herbal medicines.

The method of quality marker research and quality evaluation of traditional Chinese medicine based on drug properties and effect characteristics.

BACKGROUND: Quality of traditional Chinese medicine (TCM) plays a critical role in industry of TCM. Rapid development of TCM pharmaceutical areas is, however, greatly limited, since there are many issues not been resolved, concerning the quality study of TCM. HYPOTHESIS/PURPOSE: Core concept of TCM quality as well as the characteristics of TCM was discussed, in order to guide the quality research and evaluation of TCM, further improve the level of TCM quality control. STUDY DESIGN/METHODS: In this review, on the basis of systematic analysis of fundamental property and features of TCM in clinical application, the approaches and methods of quality marker (Q-marker) study were proposed through combination of transitivity and traceability of essentials of quality, correlation between chemical ingredients and drug property/efficacy, as well as analysis of endemicity of ingredients sharing similar pharmacophylogenetic and biosynthetic approaches. RESULTS: The approaches and methods of Q-marker study were proposed and the novel integrated pattern for quality assessment and control of TCM was established. CONCLUSION: The core concept of Q-marker has helped to break through the bottleneck of the current fragmented quality research of TCM and improved the scientificity, integrity and systematicness of quality control.

Research Advances on Hepatotoxicity of Herbal Medicines in China.

In general, herbal medicines have been considered as safe by the general public, since they are naturally occurring and have been applied in treatment for over thousands of years. As the use of herbal medicine is rapidly increasing globally, the potential toxicity of herbal drugs, in particular drug-induced liver injury (DILI), has now become a serious medical issue. According to the literature, the authors analyzed and discussed the hepatotoxicity problem of Chinese herbal medicines (CHM), including global overview on herbal-induced liver injury (HILI), current research progress on toxic CHM, diagnosis and treatment of HILI, and modern approaches and technologies of study of hepatotoxicity. As to promote the recognition of HILI and tackle the issue, a guideline for the diagnosis and treatment of HILI has recently been drafted by Chinese scientists. As suggested by the guideline, the hepatotoxicity issue of CHM, as a matter of fact, is overestimated. Up to date, the investigation of hepatotoxicity of CHM is now booming with worldwide application of CHM. This review therefore provides useful information for investigating hepatotoxicity of herbal medicine and characterizing DILI caused by CHM. In addition, authors describe in which way further efforts should be made to study the rationale of CHM and liver injury.

Challenges and Solutions of Pharmacokinetics for Efficacy and Safety of Traditional Chinese Medicine.

Traditional Chinese medicine (TCM), including herbal and folk medicine have been widely applied in healthcare field for a very long time. Based on the principle of kinetics, pharmacokinetics of TCM was developed to investigate the absorption, distribution, metabolism and excretion of active ingredients/components, single or mixed formulations, as well as the relationship between concentration and efficacy presented. Due to the complexity of TCM and its preparation, selection of molecules as markers for detection has become very difficult. So far, in which way or what indicators can be chosen for representing the pharmacodynamic action of single TCM or mixed is still controversial. Currently, a widely accepted approach is that the action of mixed preparation can be correlated by detecting pharmacokinetic behavior of one or several known active ingredients. According to the complexity of TCM, we presented a new concept of pharmacokinetic (PK) marker. The PK Marker should possess three conditions: (1) PK marker must be associated with efficacy, (2) PK marker exists in biological sample and can be determined by analytic method, and (3) PK marker should reflect the relationship between concentration and time. In TCM, four elements, "king-minister-assistant-guide", are the basic prescription principles for the individual therapy in TCM. We presented an idea of Point-line-area-cubic for four elements of TCM to study herb-herb interactions.

Drug metabolism and pharmacokinetics of nanodrugs from Chinese medicines and natural products.

Over the past few years, nanoscale Chinese medicine has become one of focuses in modern Chinese medicine research. There is an increasing need for a more systematic study on the basic issues involved in traditional Chinese medicine and a more active participation of researchers in the application area of nanoscale traditional Chinese drugs. In this review, author analyzed the current applications of nanotechnology in research and development of drugs from natural products and herbal medicines involving traditional Chinese medicines, and also discussed the bio-medicinal evaluation issues on ADME including bio-distribution and metabolism of nanodrugs. Author noted that great challenges faced in nanodrugs from herb drugs and natural products are the follows: (1) the first challenge is to prepare nanodrug delivery system and quantitatively evaluate the therapeutic effects and safety; (2) the second challenge is to clarify the concrete metabolism course; and (3) the third challenge is to study the pharmacokinetics of nanodrugs.

Herb-drug interactions involving drug metabolizing enzymes and transporters.

Herbal medicines and their active ingredients are widely used worldwide, and they have become an important part of clinical medicine. The combined use of herbs and drugs has increased the possibility of pharmacokinetic and pharmacodynamic interactions. Clinical studies have demonstrated that the combined use of herbs and drugs can enhance or attenuate the drug efficacy and toxicity. The herb-drug combinations may reduce a drug efficacy and lead to treatment failure when long-term administration. Case reports detailing serious clinical adverse reactions have promoted studies on the interactions between herbs and drugs. This review highlights recent knowledge to discuss herb-drug interactions involving metabolizing enzymes and drug transporters. Drug transporters are widely present in body and play an important role in the absorption, distribution, excretion and metabolism, efficacy, and toxicity of drugs. Investigation of transporters has developed rapidly since 1990s, the effects of many transporters on the pharmacokinetics of drugs and herb-drug interactions have been reported. Some concepts on drug transporters issued experimentally and clinically drug-drug and herb-drug interactions have applied in many studies. Methodology studies are very important for understanding the mechanism, considerations and evaluation of experiments and clinical studies on drug metabolizing enzymes and transporters in drug-drug interactions.