Phenylethylmalonamide Research Articles

High Resolution Liquid Chromatography Mass Spectrometry -A Tool for Structural Elucidation of Seed Metabolites of Abrus Precatorius(L.)

Plant metabolites with therapeutic values provide enormous scope for novel drug leads. In this context several traditional medicinal plants are targeted for phytochemicals identification and characterization worldwide. The current investigation was undertaken to identify metabolites present in seeds of Abrus precatorius (L.), a medicinal plant with potent anti-inflammatory, antimicrobial, antioxidant, antifertility, and antispermatogenic activity. Ethanol extracted seed metabolites were identified by LC-ESI-Q-TOF-MS system equipped with a G4220B pump, G4226A auto sampler and G1316C, and a diode array detector. Metabolites profiling yielded large sets of data were analyzed by METLIN Mass Spectral Database. Metabolomics analysis revealed presence of Desethyletomidate, Butabarbital, 4-aminohippurate, 20α-Dihydroprogesterone glucuronide, Phenethylamine, Etomidate, Oxyquinoline, Norfloxacin, Phenylethylmalonamide (PEMA), and Dihydro streptomycin with typical plants metabolites. The masses of these uncommon metabolites were 216.08, 212.11, 222.09, 492.27, 121.09, 244.12, 145.05, 319.12, 206.10 and 567.28 respectively. These metabolites contribute to various drug preparations used to treat human ailments.

Effects of encapsulation of primidone on its oxidative metabolism in rats.

The aim of this study was to evaluate the influence of primidone (PRM) nanoencapsulation on its metabolism. Suspensions of PRM powder and PRM-loaded poly-epsilon-caprolactone nanocapsules were administered orally in the same way to rats. Primidone-loaded poly-epsilon-caprolactone nanocapsules were prepared according to the interfacial deposition technique. Free PRM suspensions were obtained by addition of PRM powder to a suspension of 0.212% carboxymethylcellulose CMC 12H in water. The dose was 20 mg/kg, n = 6, for each experiment. Urinary and faecal levels of PRM and of its three major metabolites, phenylethylmalonamide (PEMA), phenobarbital (PB), and p-hydroxyphenobarbital (p-HO-PB), were determined. Concentrations were evaluated by high-performance liquid chromatography (HPLC) according to a validated analytical method. After PRM nanocapsule administration, non-metabolised PRM urinary levels were increased compared to those observed after administration of a suspension of primidone powder (43.7+/-8.8% after PRM-loaded nanocapsule and 37.7+/-8.1% after free PRM administration). For phenylethylmalonamide, no difference was observed in urinary excretion in the two cases. For two of the oxidised metabolites, PB and p-HO-PB, excretion was delayed and shortened. The amount of these oxidised metabolites was lowered from 0.95% after free PRM administration to 0.25% after PRM-loaded nanocapsule administration. No difference was noted in non-metabolised primidone excretion in faeces. These results suggest that primidone-loaded nanocapsules could be used as a vehicle for oral primidone administration in order to minimise the phenobarbital metabolic pathway.

Simultaneous determination of primidone and its three major metabolites in rat urine by high-performance liquid chromatography using solid-phase extraction

A new high-performance liquid chromatographic method for simultaneous determination of primidone (PRM) and of its three major metabolites, phenobarbital (PB), p-hydroxyphenobarbital ( p-HO-PB) and phenylethylmalonamide (PEMA), in rat urine, was developed. After acid hydrolysis, these compounds were extracted from urine by means of a Bond Elut Certify LRC column with good clean-up. The extracts were chromatographed on a C 1 8 reversed-phase column using isocratic elution at 40°C, with UV detection at 227 nm. The limit of detection was 0.5 mg/ml for the four compounds. Good linearity ( r 2>0.99) was observed within the calibration ranges studied: 37.4–299.3 μg/ml for PRM, 26.4–211.2 μg/ml for PB, 12.5–100.2 μg/ml for p-HO-PB and 12.1–97.0 μg/ml for PEMA. Repeatability was in the range 3.1–6.8%. This method constitutes a useful tool for studies on the influence of various parameters on primidone metabolism.

Simultaneous Determination of Primidone and Its Active Metabolites in Rat Plasma by High-Performance Liquid Chromatography Using a Solid-Phase Extraction Technique

Primidone (PRM) and its active metabolites, phenylethylmalonamide (PEMA) and phenobarbital (PB), in rat plasma were simultaneously determined using a solid‐phase extraction technique followed by high‐performance liquid chromatography (HPLC). Twenty microliters of plasma was applied to a Bond‐Elut C‐18 cartridge column with 0.1 μg of acetanilide (internal standard, IS). After the column was washed, PRM, PEMA, PB, and IS were eluted with methanol and injected into the HPLC system. Calibrations for these substances were linear in the range of 0–20 μg/mL. The coefficients of variation were 1.5–7.9% and 3.4–9.1% in the within‐day and between‐day assays, respectively. The recovery rates were 96.8–101.8%. The pharmacokinetics of these substances were examined after oral administration of PRM (50 mg/kg) to rats. TheTmaxvalues for PRM, PEMA, and PB were 1.4, 5.7, and 6.6 h, respectively, and theCmaxvalues were 18.2, 8.1, and 9.6 μg/mL, respectively. This method is useful for pharmacokinetic studies of PRM and its active metabolites.

Single-dose kinetics of primidone in human subjects: effect of phenytoin on formation and elimination of active metabolites of primidone, phenobarbital and phenylethylmalonamide.

Effect of repetitive administration of phenytoin (PHT) on the single-dose pharmacokinetics of primidone (PRM) was investigated in 3 healthy male subjects. The peak concentration of unchanged PRM was achieved at 12 and 8 h after the administration of PRM in the absence and the presence of PHT, respectively. The elimination half-life of PRM was decreased from 19.4 +/- 2.2 (mean +/- S.E.) to 10.2 +/- 5.1 h (p < 0.05) and the total body clearance was increased from 24.6 +/- 3.1 to 45.1 +/- 5.1 ml/h/kg (p < 0.01) in the presence of PHT. No significant change was observed for the apparent volume of distribution between the two treatments. In the absence of PHT, the measurable amount (> or = 0.1 mumol/l) of phenobarbital (PB) and phenylethylmalonamide (PEMA) did not appear in the serum until 5.3 and 1.3 h after the PRM administration, and the peak concentrations of PB and PEMA were achieved at 52 and 36 h, but the concentrations of both metabolites were very low (PB 1.3 mumol/l; PEMA 1.7 mumol/l). In the presence of PHT, within 0.8 and 0.5 h after the administration of PRM, the derived PB and PEMA appeared in the serum. About a 6-fold increase in the peak concentrations of both the metabolites were observed (PB 8.2 mumol/l; PEMA 11.0 mumol/l). No significant changes were observed for the elimination half-lives of both PB and PEMA in the absence and presence of PHT.(ABSTRACT TRUNCATED AT 250 WORDS)

The disposition of primidone in elderly patients.

1. The pharmacokinetics and metabolism of primidone at steady-state were studied in 10 elderly patients aged 70-81 years and eight control subjects aged 18-26 years. 2. Primidone half-lives and clearance values (mean +/- s.d.) were similar in the elderly and in the young (12.1 +/- 4.6 vs 14.7 +/- 3.5 h and 34.8 +/- 9.0 vs 33.2 +/- 7.2 ml h-1 kg-1 respectively. 3. The serum concentrations of the metabolites phenylethylmalonamide (PEMA) and phenobarbitone relative to those of parent drug were higher in the elderly than in the young, the difference being significant (P less than 0.01) in the case of PEMA. 4. The renal clearances of primidone, phenobarbitone and PEMA were moderately decreased in the elderly but this reduction was statistically significant only for PEMA. Elderly patients excreted a reduced proportion of unchanged primidone and an increased proportion of PEMA in urine. 5. Ageing is associated with a greater accumulation of PEMA, which is unlikely to have a major clinical significance.

Clearance of Phenylethylmalonamide During Haemodialysis of a Patient with Renal Failure

Information is presented for the serum concentrations during haemodialysis of primidone, phenobarbitone, and phenylethylmalonamide (PEMA) in a patient with renal failure receiving chronic primidone therapy. The concentrations of drug and metabolites fell during haemodialysis, but PEMA concentrations were above normal at all times. The average renal clearance of PEMA during 6 h of dialysis was found to be 84.7 +/- 4.6 ml min-1.

Pharmacokinetics of phenylethylmalonamide (PEMA) in elderly men.

The pharmacokinetics of phenylethylmalonamide (PEMA) were studied in 6 elderly men after oral administration of a single 400 mg dose. Peak PEMA serum levels were obtained within 4 h of intake, half-life values ranged from 30.7-57.9 h in these elderly men. The elimination half-life was twice as long when compared to a study previously performed in young volunteers.

Single-dose kinetics of primidone in acute viral hepatitis.

The pharmacokinetics of primidone (PRM) after oral administration of a single 500 mg dose was studied in 7 patients with acute viral hepatitis and 7 healthy control subjects. The elimination half-life and the apparent clearance of unchanged PRM in the patients were 18.0 +/- 3.1 h and 42 +/- 14 ml X h-1 X kg-1, respectively (mean +/- SD) and did not differ significantly from the values in the controls (half-life 17.0 +/- 2.4 h; clearance 35 +/- 8 ml X h-1 X kg-1). The metabolite phenylethylmalonamide (PEMA) was detected in the serum of all normal subjects within 2-24 h. By contrast, serum levels of this metabolite were undetectable (less than 2 mumol/1) in all but one of the patients. Serum levels of phenobarbital (PB) remained below the limit of detection (less than 2 mumol/1) in all subjects. The findings indicate that accumulation of PRM with its attendant toxicity is unlikely to occur in epileptic patients who develop acute viral hepatitis, despite evidence that the metabolism of the drug is affected by this condition. The possibility of impaired conversion to PB and its implications are discussed.

Primidone, phenobarbital, and PEMA: II. Seizure protection, neurotoxicity, and therapeutic index of varying combinations in mice.

Neurotoxicity and protection against maximal electroshock (MES) and pentylenetrazol (Metrazol) seizures were determined in mice for various combinations of primidone (PRM), phenobarbital (PB), and phenylethylmalonamide (PEMA). The results suggest that PRM and PB together are superior to either one alone in terms of spectrum of activity and relative toxicity. The protection against Metrazol and the toxicity of PB are both potentiated by PEMA at low concentrations. PEMA also potentiates the toxicity of combined PRM plus PB, without altering their protection against MES, thus lowering their therapeutic index. We conclude that PRM and PB together have an advantage over PB alone, especially when their brain concentration ratio is at or above 1 and PEMA concentrations are low. These conditions are usually not present at steady state in patients treated with PRM.

Primidone, phenobarbital, and PEMA: I. Seizure protection, neurotoxicity, and therapeutic index of individual compounds in mice.

Neurotoxicity and protection against maximal electroshock and Metrazol seizures from primidone (PRM), phenobarbital (PB), and phenylethylmalonamide (PEMA) were determined in mice for each drug separately and expressed in terms of brain concentrations. Compared with PB, PEMA was 16 times less potent against electroshock and Metrazol seizures but only 8 times less toxic. Primidone was markedly less neurotoxic than PB and equally potent against electroshock, but PRM had no effect against Metrazol or bicuculline. PRM is a relatively nontoxic anticonvulsant with a different action than PB, and PEMA is both a weak and a relatively toxic anticonvulsant.

Pharmacokinetics of phenylethylmalonamide (PEMA) after oral and intravenous administration.

The pharmacokinetics of phenylethylmalonamide (PEMA), one of the major metabolites of the antiepileptic drug primidone, have been studied in 6 healthy volunteers after administration of single 500mg intravenous and oral doses. Following intravenous administration, after a very short distributive phase (t1/2 = 0.23-0.53h), the decline of the log-PEMA concentration with respect to time appeared linear. The pharmacokinetic parameters, calculated according to a 1-compartment open model, showed the following values (mean +/- SD): terminal half-life, 15.7 +/- 3.4h; apparent volume of distribution, 0.69 +/- 0.10 L/kg; total serum clearance, 31.3 +/- 6.6 ml/h/kg. After oral administration, peak serum concentrations occurred at 0.5 to 4 hours and the oral bioavailability was 86.4 to 95.9%.

Isoniazid and Other Drugs

We read with interest the paper by Block on carbamazepine-isoniazid interaction.1 Although this report mentions that isoniazid has been previously shown to interfere with phenytoin metabolism,2 it fails to mention the interaction between isoniazid and primidone. Sutton and Kupferberg3 have described a patient in whom the addition of isoniazid prolonged the half-life of primidone (Mysoline) and decreased its biotransformation to phenobarbital and phenylethylmalonamide (PEMA). In 1964, Kato et a14 had already demonstrated the inhibitory effect of isoniazid on the metabolism of certain barbiturates in vitro.

Phenytoin: an inhibitor and inducer of primidone metabolism in an epileptic patient.

The interaction between primidone and phenytoin was studied in an epileptic patient treated with primidone only and primidone plus phenytoin for 3 months. Plasma and urine levels of drugs and metabolites were monitored daily by GC and GC-MS. The addition of phenytoin to the regimen increased steady-state plasma levels of phenobarbitone and phenylethylmalonamide (PEMA), metabolites of primidone, and decreased levels of primidone and unconjugated p-hydroxyphenobarbitone (p-OHPB), a metabolite of phenobarbitone. After withdrawal of phenytoin, plasma phenobarbitone and primidone levels slowly returned to previous steady-state levels, PEMA rapidly decreased to lower levels than before, and p-OHPB levels rose rapidly. Urinary excretion of primidone and its metabolites paralleled the changes in their plasma levels after the addition of phenytoin but the percentage of unconjugated p-OHPB in urine was unchanged during the course of the study. In conclusion phenytoin initially induces the conversion of primidone to PEMA and phenobarbitone, although each to a different extent, but it appears to inhibit the hydroxylation of phenobarbitone. Thus, two apparently contradictory phenomena seem to be involved in the primidone-phenytoin interaction. The net effect is an enhanced increase in plasma phenobarbitone levels.

Pharmacokinetics of Phenylethylmalonamide (PEMA) in Normal Subjects and in Patients Treated with Antiepileptic Drugs

The pharmacokinetics of phenylethylmalonamide (PEMA), a major metabolite of primidone, were investigated following administration of single oral doses (400 mg) to six normal subjects and six patients receiving chronic treatment with antiepileptic drugs. Peak serum PEMA levels were usually attained with 2-4 h after intake. The oral bioavailability estimated on the basis of the recovery of unchanged drug in the urine of normal subjects was at least 80%. Half-life values ranged from 17 to 25 h in normal subjects and from 10 to 23 h in the patients. No statistically significant difference in any of the calculated kinetic parameters could be found between the two groups. The data indicate that PEMA is readily absorbed from the gastrointestinal tract and that it is eliminated predominantly unchanged in the urine of man.

Single‐dose pharmacokinetics and anticonvulsant efficacy of primidone in mice

The pharmacokinetics and efficacy of the anticonvulsant primidone (PRM) and its active metabolites, phenobarbital (PB) and phenylethylmalonamide (PEMA), were studied after single-dose administration in mice. The half-life of PB is twice that of PRM and PEMA. The plasma/brain ratios provide evidence of poor penetration of PRM into brain. The results support our findings of negligible or absent PRM concentrations in the brains of patients on primidone therapy who were undergoing surgery for intractable epilepsy. The anticonvulsant properties of PRM, PB, and PEMA against maximal electroshock in mice were also studied with the use of the metabolic inhibitor SKF 525A. The half-life, potency, peak anticonvulsant effect, and effective dose curves of these compounds indicate that the anticonvulsant effect of short-term oral PRM administration in mice is from derived PB.

Simultaneous Analysis of Primidone, Phenobarbital, and Phenylethylmalonamide in Plasma and Brain

The gas-liquid chromatographic method we have developed analyzes primidone and its two active metabolites, phenobarbital and phenylethylmalonamide (PEMA), simultaneously with the use of a modified Supelco SP 2510-DA column and flame-ionization detection. Extraction and analysis are applicable to both plasma and brain samples. The drugs are quantitated underivatized. Extraction efficiency exceeded 91% for all components in both plasma and brain with coefficients of variation for primidone, phenobarbital, and PEMA of 16, 9 and 11% respectively. Mean levels for patient samples analyzed were 8.6p.g/ml for primidone, 26.0 p.g/ml for phenobarbital, and 6.2/~g/ml for PEMA. The method was used to analyze concurrent plasma and brain tissue In a kinetic study of primidone and motabolites in mice.

Synthesis of internal standards for analytical determinations of primidone, and its metabolite, phenylethylmalonamide (PEMA)

AbstractSynthetic routes are described tor the preparation of internal standards for analytical determinations of the antiepileptic agent primidone (1) and one of its primary in vivo metabolites, phenylethylmalonamide (PEMA, 3). The internal standards synthesized in the present study are p‐methylprimidone (9) and p‐methylPEMA (8). The internal standards differ in structure from the parent drug and metabolite in that the phenyl substituents of the drug and metabolite have been replaced by p‐tolyl substituents. The synthetic route for the preparation of p‐methylPEMA (8) entailed conversion of an appropriately substituted malonate to a 4,4‐disubstituted pyrazoli‐dine‐3,5‐dione, and Raney nickel reduction of the latter substance to a malonamide. This same sequence of reactions was useful for the preparation of PEMA (3) itself. A fusion of p‐methylPEMA (8) with formic acid and formamidine acetate resulted in the formation of p‐methyl‐primidone (9).

A Simultaneous High-Performance Liquid Chromatographic Analysis of the Most Common Anticonvulsants and Their Metabolites in the Serum

we present a method for the simultaneous analysis of some common anticonvulsants and their metabolites (phenylethyl malonamide, primidone, ethosuximide, 5-ethyl-5-phenylhydantoin, phenobarbital, tx-methyl-ct-phenylsuccinlmide, mephenytoin, methsuximide, mephobarbital, phenytoin, and carbamazepine) in as little as 25/JI of serum. Serum proteins are precipitated with an acetonitrile solution containing 5-(4methylphanyl)-5-phenylhydantoin, the internal standard. The drugs are eluted from a reversed-phase column with a mobile phase consisting of an acetonitrile/phosphate buffer, at a flow rate of 3.0 ml/min. The eluted drugs are detected by their absorption at 195 nm; their quantities are estimated from their peak heights. Each analysis requires no longer than 30 minutes at the optimum column temperature of 50C. The lower limit of detection for all these drugs is less than 10 ng/sample for drug standards. A sensitivity of 1.0 mglllter of serum is attained routinely for each of the drugs. Analytical recoveries for the eleven drugs varied from 88 to 102%, with good day-to-day precision (CV, between 2.0 and 8.0%). Of more than 35 drugs tested for possible Interference, only ethotoin and pentobarbital interfere with the analysis of phenobarbital and mephobarbital, respectively.

Kinetics of primidone metabolism and excretion in children

The metabolism and excretion of orally administered primidone was studied in 12 children, aged 7 to 14 yr during long-term dosing. Plasma concentrations of primidone (Pr) peaked at 4 to 6 hr and declined exponentially from 6 to 24 hr, with half-life (t1/2) values ranging from 4.5 to 11 hr. A mean of 92% (72% to 123%) of the administered dose was recovered within 24 hr from the urine as Pr and its metabolites. Of the total Pr daily dose, 42.3% (15.2% to 65.9%) was recovered as unchanged drug, 45.2% (16.3% to 65.3%) as phenylethylmalonamide (PEMA), and 4.9% (1.1% to 8.0%) as phenobarbital (Pb). The mean rate constant for conversion of Pr to PEMA (K1) was 0.0424 hr-1, for conversion of Pr to Pb (K2) was 0.0045 hr-1, and for excretion of unchanged Pr (K3) was 0.0389 hr-1. Of Pb excreted, 43% (13% to 100%) was unchanged, 15% (0% to 27%) was unconjugated p-OH Pb, 20% (0% to 44%) was conjugated p-OH Pb, and 22% (0% to 33%) was conjugated 3,4-OH Pb. KE appears to be important determinant of the steady-state plasma concentration of Pb, but interindividual differences in K2 have little influence on the overall rate constant for elimination of Pr.