Anticoagulant Phenprocoumon Research Articles

A comparative study of the molecular structure, lipophilicity, solubility, acidity, absorption and polar surface area of coumarinic anticoagulants and direct thrombin inhibitors

The methods of computational chemistry have been used to elucidate the molecular properties of coumarinic anticoagulants (acenocoumarol, phenprocoumon, warfarin and tecarfarin) and direct thrombin inhibitors (melagatran, dabigatran and their prodrug forms).

Quantifying the Effect of Covariates on Concentrations and Effects of Steady-State Phenprocoumon Using a Population Pharmacokinetic/Pharmacodynamic Model

The oral anticoagulant phenprocoumon, similar to other vitamin K antagonists, is characterized by pronounced interindividual variability in the doses needed to achieve the desired therapeutic effect. Previous studies assessed the effect of genetic and demographic covariates on empirical dose requirements of phenprocoumon to enable individualized dose prediction. The aim of the present study was to quantify major sources of interindividual variability separately on the pharmacokinetics and pharmacodynamics of phenprocoumon using a population pharmacokinetic-pharmacodynamic model. A single steady-state blood sample was collected from 278 patients and assayed by liquid chromatography-tandem mass spectrometry for phenprocoumon and its metabolites. Genotyping was performed for variants of the cytochrome P450 (CYP) 2C9 (CYP2C9) and vitamin K epoxide reductase complex, subunit 1 (VKORC1) genes. Effects were quantified by international normalized ratio (INR). Data were analyzed simultaneously using NONMEM VII. The model confirmed CYP2C9 and VKORC1 variants as the major predictors of variability in phenprocoumon concentrations and effects, together with body weight, age, comedication with CYP3A modifiers (i.e. inhibitors or inducers) and presence of atrial fibrillation. These covariates explained 50.0 % of the observed variability in the model parameters. Phenprocoumon clearance fractions mediated per CYP2C9 allele were 13.4, 9.5 and 5.7 mL/h for the 1, 2 and 3 variants, respectively. An additional clearance fraction of 5.3 mL/h was independent of CYP2C9 activity. Homozygous VKORC1 wild-type carriers were estimated to have a 2.13-fold higher phenprocoumon exposure requirement than homozygous 1173 C>T carriers to achieve the same effect on INR. The model provides a deeper insight in the separate pharmacokinetic and pharmacodynamic parts of phenprocoumon action. Thus, it provides important information for individualized dose prediction, with the option to include further covariates not studied here with known effects on individual pharmacokinetic or pharmacodynamic processes.

SUCCESSFUL TREATMENT OF CHARLES BONNET SYNDROME WITH NOCTURNAL OXYGEN SUPPLEMENTATION

To the Editor: A 92-year-old woman with visual impairment due to dry macular degeneration was admitted to the hospital in January 2004 with an 8-week history of visual hallucinations. She saw flowers growing out of the walls, animals running around, and spiders crossing her meals. These hallucinations occurred intermittently during the day and lasted for minutes to hours. Although she was fully aware that the spiders were not real, she refused to eat and had lost 5 kg of body weight. She was taking the anticoagulant phenprocoumon for atrial fibrillation, bisoprolol for heart rate control, and ramipril for hypertension. She was fit and healthy for her age, lived alone, and had been able to care for herself with only a little daily support from her 70-year-old daughter. There was no personal or family history of psychiatric problems. Neurological examination was normal except for reduced visual acuity (20/400 on the Jaeger table). Her score on the short form of the Mini-Mental State Examination was normal at 20/21. There was no evidence of mood disorder, dementia, or mental illness on psychiatric assessment. An electrocardiogram confirmed atrial fibrillation, and echocardiography showed concentric left ventricular hypertrophy with normal systolic function. Chest X-ray and magnetic resonance imaging of the brain were normal. Intoxication, delirium, substance withdrawal, neurodegenerative disorders, and psychiatric disease were excluded as the cause of her symptoms. Her hallucinations persisted over the following 10 days in spite of treatment with haloperidol (10 mg twice a day) and risperidone (2 mg twice a day). Overnight pulse oximetry revealed severe intermittent hypoxemia with an oxygen desaturation index (ODI) of 56 per hour (normal <5/hour), a lowest nocturnal arterial oxygen saturation (SaO2) of 78% (normal >89%), and 184 minutes spent with SaO2 below 90% overnight (normal 0 minutes). Polysomnography established a diagnosis of severe obstructive sleep apnea. She did not tolerate nasal continuous positive pressure ventilation. Nocturnal oxygen supplementation through nasal cannulae at a flow rate of 3 L/min led to a significant reduction in nocturnal hypoxemia (ODI 3/hour, lowest nocturnal SaO2 89%, and 3 minutes spent below SaO2 of 90% overnight). After 2 nights of oxygen treatment, the hallucinations ceased. To establish causality, oxygen administration was halted. Hallucinations recurred after 2 days and ceased when oxygen was reinstated. She was discharged home with nocturnal oxygen supplementation. When she was readmitted 3 months later for upper gastrointestinal bleeding, oxygen therapy was not given during the first few days. Vivid visual hallucinations recurred and settled again after administration of oxygen (Figure 1). The relationship between hallucinations and oxygen treatment. Complex visual hallucinations in visually impaired, psychologically normal older people characterize Charles Bonnet syndrome (CBS). It can be associated with damage at any level of the visual tract, from cataracts and retinal disease to infarction of the occipital cortex. The most widely accepted diagnostic criteria for CBS are formed complex, persistent, or repetitive stereotyped visual hallucinations; full or partial insight into the unreal nature of the perceptions; absence of hallucinations in other sensory modalities; and absence of mental disorders.1 The most effective treatment is reversal of the visual impairment. Antipsychotic drugs are commonly prescribed but are largely ineffective.2 To the authors' knowledge, this is the first report of a causal relationship between intermittent hypoxemia and CBS. Hypoxia is associated with retinal dysfunction and poor night vision.3 A possible explanation for this patient's response to oxygen treatment would be that hypoxia affected the function of her already-compromised retina. Tissue hypoxia is a pathogenetic factor in macular degeneration, causing edema and longer-lasting but reversible changes in vision.4 This could have been sufficient to precipitate the hallucinations. Intermittent hypoxemia also has an immediate and delayed effect on cognitive function through induction of inflammation and modulation of neurotransmitters.5 This may have impaired cortical compensation and adjustment to a incomplete picture received from the damaged retina, and thus precipitated CBS. This case highlights the importance of a full clinical assessment in older patients presenting with visual problems.

Direct Catalytic Benzylation of Hydroxycoumarin - Efficient Synthesis of Warfarin Derivatives and Analogues

Effective metal-catalyzed benzylations of 4-hydroxycoumarin have been developed. Employing a low amount of a cheap, nontoxic, and air-stable catalyst the 3-alkylated hydroxycoumarins were isolated in high yields after short reaction times. Applying this new methodology two widely used anticoagulants phenprocoumon and coumatetralyl were synthesized applying mild reaction conditions.

A General and Efficient Iron‐Catalyzed Benzylation of 1,3‐Dicarbonyl Compounds.

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A General and Efficient Iron‐Catalyzed Benzylation of 1,3‐Dicarbonyl Compounds

AbstractVarious CH‐acidic 1,3‐dicarbonyl compounds and methyl 3‐acetamidobut‐2‐enoate react with benzylic alcohols to give the corresponding 2‐benzylated products in good to excellent yield. Typically, reactions proceed under mild conditions (50–80 °C; air) in the presence of catalytic amounts of inexpensive iron chloride hexahydrate. The benzylation of 4‐hydroxycoumarin gives the pharmaceutically interesting 4‐hydroxy‐3‐(1‐phenylethyl)‐2H‐chromen‐2‐ones. As an example the anticoagulant Phenprocoumon is prepared in one step from commercially available substrates in 94 % yield.

Genetic polymorphisms of cytochrome P450 2C9 causing reduced phenprocoumon (S)-7-hydroxylation in vitro and in vivo

The effect of cytochrome P450 (CYP) 2C9 polymorphisms on the stereoselective biotransformation of the oral anticoagulant phenprocoumon (PPC) to inactive, monohydroxylated metabolites was studied in vitro and in vivo.In human liver microsomes, the (S)-7-hydroxylation — being the major metabolic pathway — was significantly compromised in a gene–dose-dependent manner in samples expressing the CYP2C9*2 or CYP2C9*3 allele. The CYP2C9*3/*3 genotype corresponded to an almost fourfold lower (S)-7-hydroxylation rate than CYP2C9*1/*1 (wild-type).The intrinsic clearance of human recombinant CYP2C9*2 and CYP2C9*3 for the (S)-7-hydroxylation was 28.9 and 50.9% lower than of CYP2C9*1, respectively.The area under the plasma concentration–time curve (AUC) of PPC metabolites after oral intake of 12 mg racemic PPC was significantly lower in volunteers expressing the CYP2C9*2 or CYP2C9*3 allele. Increasing plasma AUC metabolic ratios (parent compound/metabolite) in CYP2C9*2 and CYP2C9*3 variant allele carriers were found for each hydroxylation reaction and the CYP2C9*3/*3 genotype corresponded to an about 10-fold higher metabolic ratio of PPC (S)-7-hydroxylation relative to CYP2C9*1/*1.CYP2C9 polymorphisms cause a markedly compromised PPC (S)-7-hydroxylation. However, PPC metabolism appears overall less influenced by CYP2C9 genotype compared with other oral anticoagulants and it may thus be a valuable alternative for therapeutic anticoagulation of patients expressing CYP2C9 variant alleles.

Identification of cytochromes P450 2C9 and 3A4 as the major catalysts of phenprocoumon hydroxylation in vitro.

This in-vitro study aimed at an identification of cytochrome P(450) (CYP) enzymes catalysing the (S)- and (R)-hydroxylation of the widely used anticoagulant phenprocoumon (PPC) to its major, inactive metabolites. Relevant catalysts were identified by kinetic, correlation and inhibition experiments using human liver microsomes and recombinant enzymes. Kinetics revealed (S)-7-hydroxylation as quantitatively most important. Biphasic Eadie-Hofstee plots indicated more than one catalyst for the 4'-, 6- and 7-hydroxylation of both enantiomers with mean K(m1) and K(m2) of 144.5+/-34.9 and 10.0+/-6.49 microM, respectively. PPC hydroxylation rates were significantly correlated with CYP2C9 and CYP3A4 activity and expression analysing 11 different CYP-specific probes. Complete inhibition of PPC hydroxylation was achieved by combined addition of the CYP3A4-specific inhibitor triacetyloleandomycin (TAO) and a monoclonal, inhibitory antibody (mAb) directed against CYP2C8, 9, 18 and 19, except for the (R)-4'-hydroxylation that was, however, inhibited by ~80% using TAO alone. (S)-PPC hydroxylation was reduced by approximately 2/3 and approximately 1/3 using mAb2C8-9-18-19 and TAO, respectively, but (R)-6- and 7-hydroxylation by approximately 50% each. Experiments with mAbs directed against single CYP2C enzymes clearly indicated CYP2C9 as a major catalyst of the 6- and 7-hydroxylation for both enantiomers. However, CYP2C8 was equally important regarding the (S)-4'-hydroxylation. Recombinant CYP2C8 and CYP2C9 were high-affinity catalysts (K(m) <5 microM), whereas CYP3A4 operated with low affinity (K(m) >100 microM). CYP2C9 and CYP3A4 are major catalysts of (S)- and (R)-PPC hydroxylation, while CYP2C8 partly catalysed the (S)-4'-hydroxylation. Increased vigilance is warranted when PPC treatment is combined with substrates, inhibitors, or inducers of these enzymes.

Determination of phenprocoumon in plasma and urine using at-line solid-phase extraction-capillary electrophoresis

The use of capillary electrophoresis (CE) for the analysis of biological samples is rather problematic because of the large number of interferences present in the matrix. One of the possibilities to solve such problems is to couple solid-phase extraction (SPE) at-line with CE, a technique developed in our laboratory. In this study at-line SPE–CE is performed for the determination of the anticoagulant phenprocoumon in biological fluids. Plasma samples are injected after the addition of 1 vol.% of formic acid to release the drug from binding proteins, while urine samples can be directly injected. The procedure is linear between 0.2 and 30 μg ml −1 with a correlation coefficient, r 2, of 0.9996. The detection limit in plasma is 0.1 μg ml −1, which is fully adequate in view of the concentrations, that have to be dealt with in practice. The phenprocoumon concentration in a plasma sample of a patient treated with the anticoagulant was 3.8 μg ml −1.

Lack of pharmacodynamic and pharmacokinetic interaction between pantoprazole and phenprocoumon in man.

Pantoprazole is a selective proton pump inhibitor characterized by a low potential to interact with the cytochrome P450 enzymes in man. Due to the clinical importance of an interaction with anticoagulants, this study was carried out to investigate the possible influence of pantoprazole on the pharmacodynamics and pharmacokinetics of phenprocoumon. Sixteen healthy male subjects were given individually adjusted doses of phenprocoumon to reduce prothrombin time ratio (Quick method) to about 30-40% of normal within the first 5-9 days and to maintain this level. The individual maintenance doses remained unaltered from day 9 on and were administered until day 15. Additionally, on study days 11-15, pantoprazole 40 mg was given per once daily. As a pharmacodynamic parameter, the prothrombin time ratio was determined on days 9 and 10 (reference value) and on days 14 and 15 (test value), and the ratio test/reference was evaluated according to equivalence criteria. The equivalence ratio (test/reference) for prothrombin time ratio was 1.02 (90% confidence interval 0.95-1.09), thus fulfilling predetermined bioequivalence criteria (0.70-1.43). The pharmacokinetic characteristics AUC0-24h and Cmax of S(-)- and R(+)-phenprocoumon were also investigated using equivalence criteria. Equivalence ratios and confidence limits of AUC0-24h and of Cmax of S(-)-phenprocoumon (0.93, 0.87-1.00 for AUC0-24h; 0.95, 0.88-1.03 for Cmax) and of R(+)-phenprocoumon (0.89, 0.82-0.96; 0.9, 0.83-0.98) were within the accepted range of 0.8-1.25. Pantoprazole does not interact with the anticoagulant phenprocoumon on a pharmacodynamic or pharmacokinetic level. Concomitant treatment was well tolerated.

Determination of the coumarin anticoagulant phenprocoumon and metabolites in human plasma, urine and breast milk by high-performance liquid chromatography after solid-phase extraction

The coumarin anticoagulant phenprocoumon (PH) and metabolites (6-, 7- and 4′-hydroxyphenprocoumon) were analysed in plasma and urine samples from anticoagulated patients using solid-phase extraction and high-performance liquid chromatography with reversed-phase columns and ultraviolet and fluorescence detection; a simpler handling of samples, higher selectivity, precision, accuracy and analytical recovery were obtained compared to analysis using liquid—liquid extraction. Similarly, a method for the analysis of PH in human breast milk was developed to assess the passage of anticoagulants into breast milk in anticoagulated lactating women.

Direct column liquid chromatographic enantiomer separation of the coumarin anticoagulants phenprocoumon, warfarin, acenocoumarol and metabolites on an α 1-acid glycoprotein chiral stationary phase

The enantiomers of the racemic coumarin anticoagulants phenprocoumon (PH) and metabolites (4′-, 6-, 7- and 8-hydroxy-PH), warfarin (WA) and metabolites (6-, 7-hydroxy-WA and the two diastereomeric WA alcohols) and acenocoumarol were resolved by column liquid chromatography using an immobilized α 1-acid glycoprotein stationary phase; elution was performed using a phosphate buffer and isopropanol gradient with and without dimethyloctylamine as modifier, and detection by ultraviolet or fluorescence. The advantages of this method are: the procedure is simple and fast and does not require pre-column derivatization; the configuration of the enantiomers can be assigned by comparison with a reference sample with already known absolute configuration; the optical purities of these compounds can be analysed with high sensitivity; the method can be applied to the determination of the enantiomers in biological samples.

Comparative Plasma Disposition and Anticoagulant Activities of Racemic Phenprocoumon and Its Metabolites in Rats

The anticoagulant phenprocoumon (PH) and its metabolites 6-hydroxy-, 7-hydroxy- and 4'hydroxy-phenprocoumon (6-OH-PH, 7-OH-PH and 4'-OH-PH, respectively) were separately administered intravenously as racemates to rats in order to measure the potential effects of the major metabolites of PH on coagulation. Plasma samples were assayed for total concentrations of the administered compounds and the corresponding prothrombin times; kinetic parameters and anticoagulant activities were estimated using a pharmacodynamic model based on the rate of synthesis of clotting factors. The relative potencies were in the order PH > 4'-OH-PH > 6-OH-PH > 7-OH-PH the latter showing no activity. Patients on PH therapy showed lower plasma concentrations of metabolites than of PH; in humans the metabolites of PH will not contribute significantly to the overall anticoagulant activity of the drug.

Effects of the oral anticoagulant phenprocoumon on blood coagulation and thrombogenesis induced by rabbit aorta subendothelium exposed to flowing human blood: role of dose and shear rate.

We investigated the effect of oral anticoagulation on thrombogenesis induced by the subendothelium of rabbit aorta. Eighteen healthy volunteers underwent a 2-week treatment with the oral coumarin preparation phenprocoumon to a target international normalized ratio (INR) of 5. By using an ex-vivo perfusion chamber system, the interaction between flowing blood and exposed subendothelium was measured at low (50 sec-1) and high (650 sec-1) wall shear rates. The low shear rate simulated blood flow in venous vessels and the high shear rate simulated blood flow in arterial vessels. Deposition of fibrin, platelets, and platelet thrombi on subendothelium was quantified by morphometric and immunologic techniques. Fibrin deposition prevailed at the low shear rate (183 +/- 57 ng/mm2 vs 46 +/- 16 ng/mm2, low vs high shear rate; mean +/- SEM; p less than 0.01). In contrast, the interaction of platelets with subendothelium was more intense at high shear rates when compared with low shear rates, as indicated by higher platelet adhesion (44% +/- 2% vs 9% +/- 1% coverage of subendothelium with platelets, p less than 0.001) and platelet thrombus volumes (3.6 +/- 0.4 microns 3/microns 2 vs 1.3 +/- 0.3 microns 3/microns 2, p less than 0.001). Fibrin deposition on subendothelium was substantially reduced even at a low intensity of anticoagulation (reduction by 60% at INR 2.1 and by 75% at INR 3.7) and was abolished after high coumarin doses (INR 5.2). In contrast, a significant inhibition of platelet thrombus formation could be achieved only by high doses of phenprocoumon (INR 5.2). Our data indicate that relatively low doses of oral anticoagulants (INR between 2 and 3.5) substantially inhibit the fibrin formation on subendothelium prevailing at venous shear conditions, whereas a high intensity of anticoagulation (INR 4 to 5) is necessary to inhibit the formation of platelet-rich thrombi prevailing at arterial shear conditions.

Effect of oral contraceptive steroids on the pharmacokinetics of phenprocoumon.

1. The effect of chronic administration of oral contraceptive steroids (OCS) on the pharmacokinetics of the oral anticoagulant phenprocoumon was investigated in seven healthy females. 2. Plasma concentrations of phenprocoumon and the urinary recovery of unchanged as well as conjugated drug were measured following a single oral dose of 0.22 mg kg-1. A group of seven non-smoking, drug-free women matched for age and body weight served as controls. 3. Administration of OCS was associated with a significant increase in the clearance of phenprocoumon from 1.6 +/- 0.7 to 2.0 +/- 0.7 ml min-1 kg-1 (P less than 0.05). The urinary recovery of phenprocoumon glucuronide was significantly higher in OCS users (21.0 +/- 16 vs 14.0 +/- 10 (% of dose); P less than 0.05). No difference in plasma protein binding of phenprocoumon was observed, being 99.2 +/- 0.07 in both groups. 4. The accelerated glucuronidation of phenprocoumon in OCS users suggests the need for careful monitoring of the anticoagulatory response in these subjects, especially when the OCS are withdrawn.

Analysis of phenprocoumon and its hydroxylated and conjugated metabolites in human urine by high-performance liquid chromatography after solid-phase extraction

The anticoagulant phenprocoumon is mainly metabolized in humans to hydroxylated metabolites and their glucuronides. A method is described for the determination of phenprocoumon, 4′-hydroxyphenprocoumon, 6-hydroxyphenprocoumon, 7-hydroxyphenprocoumon, and their glucuronide and sulphate conjugates in human urine. Reversed-phase high-performance liquid chromatography is performed after selective extraction with disposable quaternary amine columns of untreated, and β-glucuronidase- or sulphatase-treated urine samples. Urinary excretion data are presented for total, glucuronidated, sulphated and free phenprocoumon, 4′-hydroxyphenprocoumon, 6-hydroxyphenprocoumon and 7-hydroxyphenprocoumon in twelve patients after an average daily dosage of 1.3-4.2 mg phenprocoumon.

Determination of the plasma protein binding of the coumarin anticoagulants phenprocoumon and its metabolites, warfarin and acenocoumarol, by ultrafiltration and high-performance liquid chromatography

Determination of the plasma protein binding of the coumarin anticoagulants phenprocoumon and its metabolites, warfarin and acenocoumarol, by ultrafiltration and high-performance liquid chromatography

Mass spectral analysis of phenprocoumon andO-alkyl phenprocoumons

Electron impact mass spectra of the oral coumarin anticoagulant phenprocoumon and its 4-O-alkyl and 2-O-alkyl (methyl and ethyl) derivatives (including deuterium and 13C-labelled compounds) have been analysed using accurate mass measurements and B/E and B2/E constant linked-scan techniques; fragmentation routes have been proposed which are different from those for simple coumarins and other oral coumarin anticoagulants. All compounds showed an ethyl group migration from the phenylpropyl side chain to the coumarin ring; the O-alkyl derivatives presented additionally a rearrangement with bond formation between a C atom of the O-alkyl group and the phenyl ring of the side chain. Previous literature reports about fragment ion structures for methylated phenprocoumon have been revised.

Separation of the enantiomers of phenprocoumon and warfarin by high-performance liquid chromatography using a chiral stationary phase. Determination of the enantiomeric ratio of phenprocoumon in human plasma and urine.

The enantiomers of the chiral coumarin-type anticoagulants phenprocoumon, warfarin and p-chlorophenprocoumon were separated by high-performance liquid chromatography on a chiral stationary phase (Nucleosil-Chiral 2) and normal-phase conditions. Chromatographic peak identification was performed with authentic reference compounds of the enantiomers and on-line UV spectra comparison. This method was applied to the determination of the enantiomeric ratio of phenprocoumon in plasma and urine extracts from patients under racemic drug therapy. The limit of detection (50 and 80 ng/ml) and precision (less than 5%) of the method are adequate for pharmacokinetic and enantioselective disposition studies, respectively, of phenprocoumon. No racemization was detected during the extraction procedures.

Microbial models of mammalian metabolism: production of novel alpha-diketone metabolites of warfarin and phenprocoumon using Aspergillus niger.

1. The coumarin anticoagulants warfarin and phenprocoumon were metabolized by Aspergillus niger via oxidative ring cleavage to yield the corresponding alpha-diketone metabolites. 2. Structural identification was based upon physical, spectral, and chromatographic comparisons of isolated metabolites and synthetic standards generated by the oxidative cleavage of warfarin or phenprocoumon with pyridinium chlorochromate. 3. This pathway of metabolism has been previously observed for coumarin anticoagulants in mammalian systems.