Ayurvedic preparations of Raudra Rasa inhibit agonist-mediated platelet activation and restrict thrombogenicity without affecting cell viability.

Ayurveda is considered to be one of the most ancient forms of medicine still practiced. The Ayurvedic preparation Raudra Rasa and its derivatives have been widely employed against cancer since the 12th century, but the effect of these traditional formulations on platelet function and signaling has not previously been examined. Here we demonstrate that Raudra Rasa and its derivatives significantly reduce thrombin-induced integrin activation and granule secretion in platelets, as observed by reduced PAC-1 binding and P-selectin externalization, respectively. These formulations also inhibited thrombin-stimulated phosphatidylserine exposure, mitochondrial reactive oxygen species generation, and mitochondrial transmembrane potential in platelets. Consistent with the above, Raudra Rasa significantly reduced thrombin-induced tyrosine phosphorylation of the platelet proteins, as well as phosphorylation of the enzymes AKT and GSK-3ß. In summary, Raudra Rasa inhibits agonist-mediated platelet activation without affecting cell viability, suggesting it may have therapeutic potential as an anti-platelet/anti-thrombotic agent.

An integration strategy combined progressive multivariate statistics with anticoagulant activity evaluation for screening anticoagulant quality markers in Chinese patent medicine.

ETHNOPHARMACOLOGICAL RELEVANCE: The cardiovascular and cerebrovascular diseases affect human health globally. Naoxintong capsules (NXTs), a famous Chinese Patent Medicine, has been especially applied to treat cerebral infarction and coronary heart disease in clinical practice. The anticoagulant activity of this prescription plays an important role in this course of treatment. AIM OF THE STUDY: Thrombin and factor Xa (FXa) are two key targets considering the anticoagulant activity. The purpose of this investigation is to screen the quanlity markers as key thrombin and FXa inhibitors for the anticoagulant activity oriented quality control of Chinese patent medicine. MATERIALS AND METHODS: Simple multi-polar solvent extraction processes using various proportions of solvents were conducted and their thrombin/FXa inhibitory activities were evaluated in vitro. Bivariate correlation analysis (BCA), grey correlation analysis (GCA), and orthogonal partial least squares discriminate analysis (OPLS-DA) were adopted for screening the potential active markers related to the anticoagulant activity. The chemical structures of these active compounds were identified by UHPLC-Q-TOF-MS/MS and their thrombin/FXa inhibitory activity was determined. The molecular docking technology was applied to explore the interaction between the compounds and targets. The contribution of these anticoagulant active ingredients in NXT was also investigated. Last but not the least, the contents of these markers in NXT were determined by liquid chromatography-electrospray ionization tandem triple quadrupole mass spectrometry (HPLC-ESI-MS/MS) method. RESULTS: The results showed that the NXT extract exhibited great activity against thrombin and FXa, especially extracted by 75% methanol (v/v). Six marker compounds with potential anticoagulant activity were screened out. Therein, four of the active compounds owing thrombin inhibitory activity (paeoniflorin, lithospermic acid, salvianolic acid B, Z-ligustilide) and five of the active compounds owing FXa inhibitory activity (3,5-dicaffeoylquinic acid, rosmarinic acid, lithospermic acid, salvianolic acid B and Z-ligustilide). In addition, these active compounds accounted for a large proportion of thrombin/FXa inhibitory activity of NXTs. The binding energy also showed the strong interaction formed by close connection of the compounds to the residues of targets. CONCLUSIONS: The proposed integrated stategy could be an efficient strategy to screen potential thrombin/FXa inhibitors for the bioactivity related quanlity control of Chinese patent medicine.

Screening of Potential Thrombin and Factor Xa Inhibitors from the Danshen-Chuanxiong Herbal Pair through a Spectrum-Effect Relationship Analysis.

In this study; a spectrum-effect relationship analysis combined with a high-performance liquid chromatography-mass spectrometry (LC-MS) analysis was established to screen and identify active components that can inhibit thrombin and factor Xa (THR and FXa) in Salviae Miltiorrhizae Radix et Rhizoma-Chuanxiong Rhizoma (Danshen-Chuanxiong) herbal pair. Ten potential active compounds were predicted through a canonical correlation analysis (CCA), and eight of them were tentatively identified through an LC-MS analysis. Furthermore; the enzyme inhibitory activity of six available compounds; chlorogenic acid; Z-ligustilide; caffeic acid; ferulic acid; tanshinone I and tanshinone IIA; were tested to verify the feasibility of the method. Among them; chlorogenic acid was validated to possess a good THR inhibitory activity with IC50 of 185.08 µM. Tanshinone I and tanshinone IIA are potential FXa inhibitors with IC50 of 112.59 µM and 138.19 µM; respectively. Meanwhile; molecular docking results show that tanshinone I and tanshinone IIA; which both have binding energies of less than -7.0 kcal·mol-1; can interact with FXa by forming H-bonds with residues of SER214; GLY219 and GLN192. In short; the THR and FXa inhibitors in the Danshen-Chuanxiong herbal pair have been successfully characterized through a spectrum-effect relationship analysis and an LC-MS analysis.

Based on Multi-Activity Integrated Strategy to Screening, Characterization and Quantification of Bioactive Compounds from Red Wine.

According to French Paradox, red wine was famous for the potential effects on coronary heart disease (CHD), but the specific compounds against CHD were unclear. Therefore, screening and characterization of bioactive compounds from red wine was extremely necessary. In this paper, the multi-activity integrated strategy was developed and validated to screen, identify and quantify active compounds from red wine by using ultra high performance liquid chromatography-fraction collector (UHPLC-FC), ultra fast liquid chromatography-quadrupole-time-of-flight/mass spectrometry (UFLC-Q-TOF/MS) and bioactive analysis. UHPLC-FC was employed to separate and collect the components from red wine, which was further identified by UFLC-Q-TOF/MS to acquire their structural information. Furthermore, the active fractions were tested for antioxidant activity, inhibitory activity against thrombin and lipase activities in vitro by the activity screening kit. As the results, there were 37 fractions had antioxidant activity, 22 fractions had thrombin inhibitory activity and 28 fractions had lipase inhibitory activity. Finally, 77 active components from red wine were screened and 12 ingredients out of them were selected for quantification based on the integration of multi-activity. Collectively, the multi-activity integrated strategy was helpful for the rapid and effective discovery of bioactive components, which provided reference for exploring the health care function of food.

Thrombin-based discovery strategy of bioactive-chemical quality marker combination for pollen of Typha orientalis by metabolomics coupled with chemometrics.

BACKGROUND: It is of utmost significance to choose the bioactive components as quality markers for ensuring the effectiveness of traditional Chinese medicine (TCM). Nonetheless, some markers are able to assess effectively the quality of TCM without considering the pharmacological mechanisms and intrinsic chemical complexities. OBJECTIVE: This underscores the need to discover new and efficient markers which can assess both quality and mechanism of action. Herein, a strategy of bioactive-chemical quality marker combination was proposed to improve the level of the quality control of TCM by metabolomics coupled with chemometrics. METHODS: A four-step plan was followed. Firstly, acquisition of metabolic features and component characterization of different batches of pollen of Typha orientalis C.Presl were performed using UHPLC-Q-TOF/MS. Secondly, the direct inhibitory effects of pollen of T. orientalis on thrombin was assessed by using chromogenic substrate method together with HPLC. Thereafter, bioactive-chemical marker combination associated with anti-thrombin segregation was screened using supervised classifiers. Finally, quantitative assay and prediction-model of selected markers were established for guarantying the quality of pollen of T. orientalis. RESULTS: A total of 22 compounds were annotated based on comparison with previous work from pollen of T. orientalis by UHPLC-Q-TOF/MS. Citric acid and linolenic acid inhibited the thrombin activity with IC50 values, 0.52 ± 0.02 and 0.51 ± 0.02 mg/mL, respectively. A bioactive-chemical marker combination including citric acid, linolenic acid, typhaneoside, and isorhamnetin-3-O-neohesperidoside were discovered and selected as quality markers for evaluation of pollen of T. orientalis according to their capacity for inhibiting thrombin. CONCLUSION: The thrombin-based discovery strategy of bioactive-chemical marker combination was a powerful tool for screening the quality markers for evaluation of pollen of T. orientalis.

The hirudin-like factors HLF3 and HLF4-hidden hirudins of European medicinal leeches.

The hirudin-like factors 3 (HLF3) and 4 (HLF4) belong to a new class of leech-derived factors and are present in specimens of the three European medicinal leeches, Hirudo medicinalis, Hirudo verbana, and Hirudo orientalis, respectively. Here we describe the functional analysis of natural and synthetic variants of HLF3 and HLF4. Whereas the natural variants display only very low or no detectable anti-coagulatory activities, modifications within the N-termini in combination with an exchange of the central globular domain have the potency to greatly enhance the inhibitory effects of respective HLF3 and HLF4 variants on blood coagulation. Our results support previous observations on the crucial importance of all parts (both the N- and C-termini as well as the central globular domains) of hirudin and HLF molecules for thrombin inhibition.

An online dual-enzyme co-immobilized microreactor based on capillary electrophoresis for enzyme kinetics assays and screening of dual-target inhibitors against thrombin and factor Xa.

In this study, an online capillary electrophoresis (CE) based dual-enzyme (thrombin and factor Xa) co-immobilized microreactor (THR-FXa IMER) was constructed for studying enzyme kinetics and screening dual-target inhibitors against THR and FXa with the aid of the polydopamine/graphene oxide (PDA/GO) coating. Based on the developed THR-FXa IMER, the Michaelis-Menten constants (Km) of THR and FXa were calculated to be 187.26 and 48.80 µM, respectively. The inhibition constants (Ki) for two known inhibitors, argatroban and rivaroxaban, on THR and FXa were determined to be 14.73 and 0.41 nM, respectively. In addition, after 30 consecutive runs, the enzymes' activity was remained 98% of the initial immobilized activity for both THR and FXa, which shows that the constructed IMER has good stability and repeatability. Finally, the developed method was successfully applied to screen dual-target inhibitors against THR and FXa from 30 small molecular compounds. Among them, 10 compounds such as salvianolic acid C and epigallocatechin gallate (EGCG) have dual-enzyme inhibitory activity, and 2 compounds named saikosaponin A and oleuropein have single THR inhibitory activity, 5 compounds such as rosemary acid and salvianolic acid B have single FXa inhibitory activity. Finally, the molecular interactions between enzyme and potential inhibitors were further verified via the molecular docking, and a new compound with a theoretically good coagulation inhibition effect was designed by the scaffold hopping study. In summary, the developed THR-FXa IMER is a reliable method for screening THR and/or FXa inhibitors.

Rivaroxaban and Dabigatran for Suppression of Mechanical Heart Valve-Induced Thrombin Generation.

BACKGROUND: Patients with mechanical heart valves (MHVs) require warfarin to prevent thromboembolism. Dabigatran was less effective than warfarin in patients with MHVs, which prompted a black box warning against the use of direct oral anticoagulants for this indication. However, rivaroxaban and apixaban, which inhibit factor Xa, have not been evaluated in patients with MHVs. To determine whether rivaroxaban and apixaban would be effective, we used MHV-induced thrombin generation assays to compare them with warfarin either alone or in combination with dabigatran. METHODS: Thrombin generation in the absence or presence of MHV leaflets or sewing ring segments (SRSs) was quantified. Studies were done in control plasma; plasma from patients on warfarin; plasma containing varying concentrations of rivaroxaban, apixaban, or dabigatran alone; or plasma containing rivaroxaban plus dabigatran. RESULTS: Mean endogenous thrombin potential (ETP) increased 1.2-fold, 1.5-fold, and 1.8-fold in the presence of leaflets, Teflon (Terumo Aortic (Sunrise, FL)) SRSs, or Dacron (Terumo Aortic (Sunrise, FL)) SRSs, respectively. Rivaroxaban and apixaban reduced ETP at concentrations above 50 ng/mL but were less effective than warfarin. When rivaroxaban and dabigatran were combined, they suppressed ETP in a more than additive manner. CONCLUSIONS: Whereas warfarin suppresses MHV-induced thrombin generation, MHVs induce the generation of factor Xa in concentrations that overwhelm clinically relevant concentrations of rivaroxaban or apixaban. When used in combination, rivaroxaban and dabigatran are more effective than either agent is alone, suggesting that concomitant inhibition of factor Xa and thrombin is better than inhibition of either clotting enzyme alone.

Biflavones from Ginkgo biloba as inhibitors of human thrombin.

Ginkgo Biloba leaf extract has been widely used for the prevention and treatment of thrombosis and cardiovascular disease in both eastern and western countries, but the bioactive constituents and the underlying mechanism of anti-thrombosis have not been fully characterized. The purpose of this study was to investigate the inhibitory effects of major constituents in Ginkgo biloba on human thrombin, a key serine protease regulating the blood coagulation cascade and the processes of thrombosis. To this end, a fluorescence-based biochemical assay was used to assay the inhibitory effects of sixteen major constituents from Ginkgo biloba on human thrombin. Among all tested natural compounds, four biflavones (ginkgetin, isoginkgetin, bilobetin and amentoflavone), and five flavonoids (luteolin, apigenin, quercetin, kaempferol and isorhamnetin) were found with thrombin inhibition activity, with the IC50 values ranging from 8.05 µM to 82.08 µM. Inhibition kinetic analyses demonstrated that four biflavones were mixed inhibitors against thrombin-mediated Z-GGRAMC acetate hydrolysis, with the Ki values ranging from 4.12 µM to 11.01 µM. Molecular docking method showed that the four biflavones could occupy the active cavity with strong interactions of salt bridges and hydrogen bonds. In addition, mass spectrometry-based lysine labeling reactivity assay suggested that the biflavones could bind on human thrombin at exosite I rather than exosite II. All these findings suggested that the biflavones in Ginkgo biloba were naturally occurring inhibitors of human thrombin, and these compounds could be used as lead compounds for the development of novel thrombin inhibitors with improved efficacy and high safety profiles.

Mass Spectrometry-Sensitive Probes Coupled with Direct Analysis in Real Time for Simultaneous Sensing of Chemical and Biological Properties of Botanical Drugs.

The development of botanical materials as therapeutic agents involves the meticulous assessment of safety, efficacy, and quality. Compared with small-molecule drugs, quality control of botanical drugs confronts with more significant challenges due to their inherent complexity. Current quality control methods for botanical drugs, either prevailing chemical tests or emerging biological assays, are not able to meet recent demands of multiplexing, sensitivity, and speed. Here, we propose an on-demand strategy based on a direct analysis in real time-mass spectrometry (DART-MS) platform, which is capable of simultaneously analyzing multiple constituents and bioactivities of botanical drugs. Notably, the bioactivities are assessed by a multiple-enzyme assay that adopts cleavable mass spectrometry probes as enzymatic substrates: these probes labeled with a piperazine tag make possible sensitive, multiplexed, and quantitative enzyme activity measurements. The concept is successfully demonstrated via a case study of Danshen (Salvia miltiorrhiza) Injection where simultaneous detection of 34 constituents and inhibitory activities on two target enzymes can be achieved in just minutes. This proof-of-concept application also gives evidence that combining MS-sensitive probes with DART-MS can provide an environmentally friendly, highly sensitive analytical approach for botanical quality control.

Hirudins of the Asian medicinal leech, Hirudinaria manillensis: same same, but different.

Blood coagulation in vertebrates is a complex mechanism that involves the precisely coordinated and regulated action of a cascade of factors in order to prevent excessive blood loss upon wounding. Any blood sucking ectoparasite, however, has to circumvent this mechanism to ensure the uptake of an adequate blood meal. Inhibitors of blood coagulation in the saliva are hence widespread among these animals. Thrombin as a key factor of blood coagulation is a prominent target of such inhibitors, and hirudin is probably the best known among the thrombin inhibitors. Hirudin was originally described in the genus Hirudo, but occurs in other leech genera like Hirudinaria and Macrobdella as well. Besides several isoforms of hirudin, a new class of putative leech saliva components, the hirudin-like factors (HLFs), was identified in both genera Hirudo and Hirudinaria. Here, we describe the expression, purification, and functional characterization of three HLFs (HLF5, 6, and 8, respectively) and two additional hirudins (HM3 and HM4) of Hirudinaria manillensis. While HLF6 lacked any inhibitory activity on thrombin, HLF5 as well as HLF8 clearly exhibited anticoagulatory properties. The inhibitory activity of HLF5 and HLF8, however, was much lower compared with both HM3 and HM4 of Hirudinaria manillensis as well as the hirudin variants 1 (HV1) and 2 (HV2) of Hirudo medicinalis. Neither an inhibition of trypsin nor a platelet aggregation was caused by HLF8. Our data indicates the presence of two classes (rather than isoforms) of hirudins in Hirudinaria manillensis with markedly different inhibitory activity on human thrombin.

Thrombin Inhibition: Preliminary Assessment of the Anticoagulant Potential of Turnera subulata (Passifloraceae).

Cardiovascular and thromboembolic disturbances are the main causes of disease-related deaths worldwide. Regardless of the etiological factors involved in thrombus formation, coagulation is mainly activated by thrombin, one of the most important blood clotting molecules. Thus, this study evaluated the Turnera subulata leaf crude extract, its ethyl acetate fraction effect on the coagulation cascade, and its possible side effects. Their phytocomposition indicated polyphenols, mainly flavonol-3-O-glycosylate and a flavone glycoside, without in vitro and in vivo toxicity. Regarding their potential anticoagulants, results displayed partial thromboplastin and prothrombin time activation, and Xa and IIa, and thrombin inhibition by heparin II cofactor, indicating significant anticoagulant activity, suggesting direct and indirect thrombin inhibition as the main mechanism of action. Therefore, T. subulata leaf active compounds exhibit therapeutic potential required to develop phytotherapeutic formulations to assist conventional anticoagulants in clinical treatments.

The Non-Vitamin K Antagonist Oral Anticoagulants in Heart Disease: Section V-Special Situations.

Non-vitamin K antagonist oral anticoagulants (NOACs) include dabigatran, which inhibits thrombin, and apixaban, betrixaban, edoxaban and rivaroxaban, which inhibit factor Xa. In large clinical trials comparing the NOACs with the vitamin K antagonist (VKA) warfarin, dabigatran, apixaban, rivaroxaban and edoxaban were at least as effective for stroke prevention in atrial fibrillation and for treatment of venous thromboembolism, but were associated with less intracranial bleeding. In addition, the NOACs are more convenient to administer than VKAs because they can be given in fixed doses without routine coagulation monitoring. Consequently, the NOACs are now replacing VKAs for these indications, and their use is increasing. Although, as a class, the NOACs have a favourable benefit-risk profile compared with VKAs, choosing among them is complicated because they have not been compared in head-to-head trials. Therefore, selection depends on the results of the individual trials, renal function, the potential for drug-drug interactions and preference for once- or twice-daily dosing. In addition, several 'special situations' were not adequately studied in the dedicated clinical trials. For these situations, knowledge of the unique pharmacological features of the various NOACs and judicious cross-trial comparison can help inform prescription choices. The purpose of this position article is therefore to help clinicians choose the right anticoagulant for the right patient at the right dose by reviewing a variety of special situations not widely studied in clinical trials.

Anti-Thrombin, Anti-Adhesive, Anti-Migratory, and Anti-Proliferative Activities of Sulfated Galactans from the Tropical Green Seaweed, Udotea flabellum.

In this study, sulfated polysaccharide-rich extracts were isolated from 22 tropical seaweeds (4 red, 11 brown, and 7 green) found in northeastern Brazil, and evaluated for the role of anticoagulant agents. Fifteen of the extracts showed anticoagulant activity, including all the extracts from green seaweeds. Udotea flabellum (a green seaweed) extract was the most potent, requiring an amount of only 3 µg to double the plasma coagulation time in the activated partial thromboplastin time test. A similar result was obtained with 1 µg of heparin. Two sulfated homogalactans with anticoagulant activity, F-I (130 kDa) and F-II (75 kDa), were isolated from this extract using several bio-guided purification steps. Their anticoagulant activity, as well as properties related to antitumor activity (anti-proliferative, anti-adhesive, and anti-migratory), were accessed. Their anticoagulant activities were close to that of heparin. We found that F-I and F-II (0.5⁻10 µg/mL) were not able to directly inhibit thrombin. In the presence of anti-thrombin, F-I (0.5 µg/mL) was more effective than heparin (0.5 µg/mL) in inhibiting thrombin, while F-II showed similar effects as heparin. F-I and F-II also inhibited B16-F10 (murine melanoma cells) adhesion, migration, and proliferation on a fibronectin-coated surface, but not on laminin- or collagen I-coated surfaces. Except for the antiproliferative activity, the other effects of F-I and F-II were eliminated upon their desulfation (~50%), indicating that the degree of sulfation is not as important for F-I and F-II anti-proliferative activity as the sulfation position. Taken together, the results provide strong evidence for the potential utility of sulfated galactans from U. flabellum, making these compounds an interesting option for future investigations that aim to design new anticoagulant/antitumor agents.

[Quality evaluation of musk based on the biological potency of its anti-thrombin activty].

This study attempts to establish a method for the anti-thrombin activty bioassay of musk, explore the impact of species and producing areas on the anti-thrombin activty of musk, and provide scientific basis for its biological quality evaluation. Anti-thrombin activty of musk was analyzed by thrombin titration, and the influence factors such as musk solution concentration, fibrinogen concentration, thrombin concentration and titration interval were optimized to evaluate the effect of different species and producing areas on anti-thrombin activty of musk. As a result, there was a good linear relationship between musk solution concentration and thrombin consumption volume within the range of 0.01-0.02 g·mL⁻¹ (r=0.991 4) under the experimental condition as follows: fibrinogen concentration was 0.5%, the thrombin concentration was 10 U·mL⁻¹ï¼› titration time interval was once every minute, and each titration volume was 2 µL. The average anti-thrombin activty potency of Moschus berezovskii from different producing areas was (105.0±10.4) U·g⁻¹, (102.4±5.5) U·g⁻¹ for M. sifanicus from different producing areas, (97.7±6.6) U·g⁻¹ for M. moschiferus from Anhui province, and (58.6±6.4) U·g⁻¹ for artificial musk. The results indicated that this anti-thrombin activty bioassay method could be applied to evaluate the anti-thrombin activty of musk quickly, conveniently, sensitively and exactly. It was also suggested that different species and producing areas had effects on the anti-thrombin activty of musk, so it is necessary to pay attention to species and producing areas in the process of musk domestication; meanwhile, the artificial musk and natural musk also showed great differences in anti-thrombin activty, suggesting that ratio in artificial musk prescription needs to be further optimized.

Evaluation of thrombin inhibitory activity of catechins by online capillary electrophoresis-based immobilized enzyme microreactor and molecular docking.

An online capillary electrophoresis (CE)-based thrombin (THR) immobilized enzyme microreactor (IMER) method was established to screen THR inhibitors in this study. S-2366 was used as chromogenic substrate for determination of THR activity and other kinetic constants. After continuously run for 50 times, the prepared IMER could still remain 89% of the initial immobilized enzyme activity. The Michaelis-Menten constant (Km) of immobilized THR was measured as 0.514 mmol/L and the half-maximal inhibitory concentration (IC50) and inhibition constant (Ki) of argatroban on THR were determined as 78.07 and 26.53 nmol/L, respectively, which indicated that CE-based THR IMER was successfully established and could be applied to screen THR inhibitors. Then the prepared IMER was used to investigate the inhibitory potency on THR of four main catechins in green tea including epicatechin (EC), epigallocatechin (EGC), epicatechin gallate (ECG), and epigallocatechin gallate (EGCG). The results showed that ECG and EGCG had good THR inhibition activity and their inhibition rates at concentration of 200 µmol/L were 53.2 ±â€¯3.8% and 55.8 ±â€¯2.6%, respectively, which was in consistent with the results of microplate reader assay. Additionally, molecular docking results showed that the benzopyran groups of ECG and EGCG were inserted into the THR active pocket and interacted with residues LYS60F, TRP60D, TRY60A, IEU99, GLY216, HIS57 and SER195, but EC and EGC did not. Therefore, the developed CE-based THR IMER is reliable method for measuring THR inhibitory activity of natural inhibitors.

Effect of Xylan Sulfates on Coagulation of Human Blood Plasma.

Sulfated derivatives of xylan (isolated from Bétula pubéscens wood) with average molecular weight ~34 kDa, sulfur content of 11.3-17.5%, a degree of substitution of 0.74-1.64 are anticoagulants of direct type of action. Antithrombin and antifactor Xa activities in three tested xylan samples did not differ and reached 30.8-31.8 and 13.5-14.3 U/mg, respectively.

Inhibitory Effect of Triterpenoids from Panax ginseng on Coagulation Factor X.

Enzymes involved in the coagulation process have received great attention as potential targets for the development of oral anti-coagulants. Among these enzymes, coagulation factor Xa (FXa) has remained the center of attention in the last decade. In this study, 16 ginsenosides and two sapogenins were isolated, identified and quantified. To determine the inhibitory potential on FXa, the chromogenic substrates method was used. The assay suggested that compounds 5, 13 and 18 were mainly responsible for the anti-coagulant effect. Furthermore, these three compounds also possessed high thrombin selectivity in the thrombin inhibition assay. Furthermore, Glide XP from Schrödinger was employed for molecular docking to clarify the interaction between the bioactive compounds and FXa. Therefore, the chemical and biological results indicate that compounds 5 (ginsenoside Rg2), 13 (ginsenoside Rg3) and 18 (protopanaxtriol, PPT) are potential natural inhibitors against FXa.

[Non-vitamin K dependent oral anticoagulants : What is important in intensive care medicine]. / Nicht-Vitamin-K-abhängige orale Antikoagulanzien : Was ist in der Intensivmedizin zu beachten.

Since first used in 2009, non-vitamin K oral anticoagulants (NOAC) have gained world-wide acceptance. Two groups of NOAC are currently used: the direct thrombin antagonist dabigatran and three direct factor  Xa antagonists apixaban, edoxaban, and ricaroxaban. With their increasing use for prevention of thromboembolism, the probability increases that NOAC-pretreated patients are admitted to emergency departments or intensive care units.The clinical challenge in NOAC preanticoagulated patients is to adequately cope with the given anticoagulated status of such patients. Because of their short half-life, many patients will be adequately treated with a "wait and see" approach, and surgeries and interventions are postponed until anticoagulant activities have totally subsided. In the few cases where immediate action is mandated, based on appropriate risk assessments it can be decided either to take the increased hemorrhagic risk of early intervention or to transfuse factor concentrates like PPSB or FEIBA which can safely reverse the anticoagulant activities of the three factor Xa antagonists (and potentially also of dabigatran). Recently a humanized Fab antibody fragment for dabigatran, idarucizumab, has been introduced onto the market, that can immediately reverse the anticoagulant effects of dabigatran. For the reversal of dabigatran, idarucizumab is therefore the drug of choice.In addition, in some specific indications of emergency and intensive care medicine, the primary use of a NOAC can be considered advantageous. Such indications are early cardioversion in patients admitted for new episodes of atrial fibrillation and patients with acute pulmonary embolism. For the widespread use of low-molecular-weight heparins in such indications, however, the decision to use a NOAC for anticoagulant therapy is frequently postponed to the treatment phase when the stabilized patient is already treated on the general ward.

Screening Explorer-An Interactive Tool for the Analysis of Screening Results.

Screening Explorer is a web-based application that allows for an intuitive evaluation of the results of screening experiments using complementary metrics in the field. The usual evaluation of screening results implies the separate generation and apprehension of the ROC, predictiveness, and enrichment curves and their global metrics. Similarly, partial metrics need to be calculated repeatedly for different fractions of a data set and there exists no handy tool that allows reading partial metrics simultaneously on different charts. For a deeper understanding of the results of screening experiments, we rendered their analysis straightforward by linking these metrics interactively in an interactive usable web-based application. We also implemented simple consensus scoring methods based on scores normalization, standardization (z-scores), and compounds ranking to evaluate the enrichments that can be expected through methods combination. Two demonstration data sets allow the users to easily apprehend the functions of this tool that can be applied to the analysis of virtual and experimental screening results. Screening Explorer is freely accessible at http://stats.drugdesign.fr .