Pharmacokinetics Of Valproic Acid Research Articles

A rapid and highly sensitive UPLC–MS/MS method using pre-column derivatization with 2-picolylamine for intravenous and percutaneous pharmacokinetics of valproic acid in rats

A rapid, highly sensitive and specific ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) for the detection of valproic acid (VPA) in rat plasma following the topical application was developed and validated. This method was carried out with pre-column derivatization using 2-picolylamine (PA) which reacts with the carboxylic acid group of VPA. The derivatization was completed in 10min and the resulting PA-VPA derivative enabled the sensitive detection of VPA in selected reaction monitoring (SRM) mode. Sample preparation was done with simple liquid-liquid extraction and chromatographic separation was achieved within 5min on a C18 column using a gradient elution with the mobile phase of 2mM ammonium formate containing 0.1% formic acid and methanol. The standard curves were linear over the concentration range of 0.07-200μg/mL with a correlation coefficient higher than 0.99. The limit of detection (LOD) and the lower limit of quantification (LLOQ) was 0.03 and 0.07μg/mL, respectively with 100μL of plasma sample. The intra- and inter-day precisions were measured to be below 10.7% and accuracies were within the range of 94.1-115.9%. The validated method was successfully applied to the pharmacokinetics of VPA in the rat following topical and intravenous applications.

Whole-body pharmacokinetics of HDAC inhibitor drugs, butyric acid, valproic acid and 4-phenylbutyric acid measured with carbon-11 labeled analogs by PET

The fatty acids, n-butyric acid (BA), 4-phenylbutyric acid (PBA) and valproic acid (VPA, 2-propylpentanoic acid) have been used for many years in the treatment of a variety of CNS and peripheral organ diseases including cancer. New information that these drugs alter epigenetic processes through their inhibition of histone deacetylases (HDACs) has renewed interest in their biodistribution and pharmacokinetics and the relationship of these properties to their therapeutic and side effect profiles. In order to determine the pharmacokinetics and biodistribution of these drugs in primates, we synthesized their carbon-11 labeled analogues and performed dynamic positron emission tomography (PET) in six female baboons over 90 min. The carbon-11 labeled carboxylic acids were prepared by using (11)CO2 and the appropriate Grignard reagents. [(11)C]BA was metabolized rapidly (only 20% of the total carbon-11 in plasma was parent compound at 5 min post injection) whereas for VPA and PBA 98% and 85% of the radioactivity were the unmetabolized compound at 30 min after their administration respectively. The brain uptake of all three carboxylic acids was very low (<0.006%ID/cc, BA>VPA>PBA), which is consistent with the need for very high doses for therapeutic efficacy. Most of the radioactivity was excreted through the kidneys and accumulated in the bladder. However, the organ biodistribution between the drugs differed. [(11)C]BA showed relatively high uptake in spleen and pancreas whereas [(11)C]PBA showed high uptake in liver and heart. Notably, [(11)C]VPA showed exceptionally high heart uptake possibly due to its involvement in lipid metabolism. The unique biodistribution of each of these drugs may be of relevance in understanding their therapeutic and side effect profile including their teratogenic effects.

Sex related differences on valproic acid pharmacokinetics after oral single dose

Plasma concentration-time data obtained after an oral single dose of 500-mg valproic acid (VPA) in a delayed release formulation was used to model enterohepatic cycling of the drug. Fourteen healthy subjects (seven women and seven men) were enrolled, food intake was standardized, blood samples were withdrawn up to 48h post dosing and VPA plasma concentrations were analyzed by HPLC-UV method. Secondary peaks of VPA were observed in almost all subjects. A population pharmacokinetic analysis with sex, weight, age and contraceptive therapy as possible covariates was performed. Women without contraceptive therapy presented a longer lag time, a lower VPA hepatic output and a higher reabsorbed fraction than men. Weight affected both volume of the central compartment and the absorption rate constant. Women under contraceptive therapy showed similar disposition parameters to men. Reabsorbed fraction of bioavailable dose was 46.2% in women and 21.8% in men, revealing important sex differences in drug hepatobiliar output. These results showed that VPA displays sex-related pharmacokinetic differences due to different metabolic and transporter-mediated disposition.

The Effect of Uridine Diphosphate Glucuronosyltransferase (UGT)1A6 Genetic Polymorphism on Valproic Acid Pharmacokinetics in Indian Patients with Epilepsy: A Pharmacogenetic Approach

Sodium valproate is a widely prescribed broad-spectrum antiepileptic drug. It shows high inter-individual variability in pharmacokinetics and pharmacodynamics and has a narrow therapeutic range. We evaluated the effects of polymorphic uridine diphosphate glucuronosyltransferase (UGT)1A6 (541A>G, 552A>C) metabolizing enzyme on the pharmacokinetics of sodium valproate in the patients with epilepsy who showed toxicity to therapy. Genotype analysis of the patients was made with polymerase chain-restriction fragment length polymorphism (RFLP) with sequencing. Plasma drug concentrations were measured with reversed phase high-performance liquid chromatography (HPLC) and concentration-time data were analyzed by using a non-compartmental approach. The results of this study suggested a significant genotypic as well as allelic association with valproic acid toxicity for UGT1A6 (541A>G) or UGT1A6 (552A>C) polymorphic enzymes. The elimination half-life (t ½=40.2h) of valproic acid was longer and the clearance rate (CL=917ml/h) was lower in the poor metabolizers group of UGT1A6 (552A>C) polymorphism who showed toxicity than in the intermediate metabolizers group (t ½=35.5h, CL=1,022ml/h) or the extensive metabolizers group (t ½=25.4h, CL=1,404ml/h). Our findings suggest that the UGT1A6 (552A>C) genetic polymorphism plays a significant role in the steady state concentration of valproic acid, and it thereby has an impact on the toxicity of the valproic acid used in the patients with epilepsy.

Circadian variation of Valproic acid pharmacokinetics in mice

Valproic acid (VPA) is currently one of the most commonly used antiepileptic drugs. This study aims to investigate whether VPA pharmacokinetics varied according to circadian dosing-time. A single dose of VPA (350mgkg−1) was administered by intraperitonally (i.p.) route to a total of 132 mice synchronized for 3weeks to 12h light (rest span) and 12h dark (activity span). Four different circadian times (1, 7, 13 and 19 HALO) of drug injection were used (33 mice/circadian time). At each circadian time, blood samples were withdrawn at (0h) and at 0.083, 0.166, 0.25, 0.5, 0.75, 1, 1.25, 1.5, 2 and 3h after VPA injection. Plasma VPA concentrations were determined by an EMIT method.There were no significant differences in Tmax of VPA whatever the circadian-time of injections (Tmax=0.166h). However, there were relevant differences in Cmax between the four circadian groups (p<0.005), it varied between 386±30.86mgL−1 in mice treated at 7 HALO and 824±39.85mgL−1 in mice treated at 19 HALO. The AUC0–∞ was significantly two times higher when VPA was administered at 19 HALO as compared to the injection at 7 HALO. Drug dosing at 7 HALO resulted in highest ClT value: 0.405±0.006Lh−1kg−1, whereas ClT was significantly slower when VPA was administered at 19 HALO (0.157±0.009Lh−1kg−1) (p<0.0001). The AUC0–∞ was significantly 2-fold higher when VPA was administered at 19 HALO (2216.65±138.91mgh−1L−1) as compared to the injection at 7 HALO (864.09±16.82mgh−1L−1) (p<0.0001).Cosinor analysis showed circadian rhythm in different pharmacokinetic parameters. Cmax and AUC0–∞ have a significant circadian rhythm with an acrophase located at 20.16 HALO±0.16h (the middle of the active span) (p<0.001), whereas ClT and Vd showed a significant circadian rhythm with an acrophase located respectively at 7.86 HALO±0.57h and 6.13 HALO±0.07h (the middle of the rest span) (p<0.001).The large circadian variation of VPA pharmacokinetic processes might be involved in the mechanisms of circadian rhythm in murine toxicity since the optimal tolerance corresponded to the time which induces lowest Cmax and AUC values.

Valproic acid pathway

Valproic acid (VPA) is a branched short-chain fatty acid derived from naturally occurring valeric acid. VPA is used primarily in the treatment of epilepsy and seizures, but is also used in migraine, bipolar, mood, anxiety, and psychiatric disorders [1]. Recent work has explored its use as an adjuvant agent in cancer, HIV therapy, and neurodegenerative disease because of its action as histone deacetylase (HDAC) inhibitor [2]. VPA is widely used in pediatric epilepsy because of its multiple mechanisms of action and acceptable safety profile [3]. The dose requirements for VPA are highly variable (10-fold differences in mean dose in adults) [4] and interactions with other drugs are common (discussed below). Therapeutic drug monitoring is commonly used, although both the clinical and toxic effects of the drug are considered to be poorly correlated with total serum concentrations [3]. The drug label carries a black box warning for life-threatening adverse drug reactions (ADR) including hepatoxocity, teratogenicity, and pancreatitis [1]. Compared with adults, children appear to be at an increased risk for severe hepatotoxic reactions to VPA, in particular those younger than 2 years of age undergoing polytherapy, with existing developmental delays and coincident metabolic disorders [1]. Hyperammonemia is also a documented ADR of VPA treatment, although this is usually successfully resolved by cessation of VPA and treatment with carnitine [5]. The Food and Drug Administration has issued recommendations for patient testing and advises that VPA is contraindicated in patients with known urea cycle disorders (see Clinical PGx tab at http://www.pharmgkb.org/drug/PA451846). However, specific genetic tests for the determination of at-risk patients are not mentioned. The aim of this study was to introduce candidate genes in the pharmacokinetics (PK) (Fig. 1), pharmacodynamics (PD) of VPA (Fig. 2), and discuss results from pharmacogenomic studies so far. Fig. 1 Graphic representation of the candidate genes involved in valproic acid (VPA) pharmacokinetics. A fully interactive version of this pathway is available online at PharmGKB at http://www.pharmgkb.org/pathway/PA165964265. CYP, cytochrome P450. Fig. 2 Graphic representation of the candidate genes involved in valproic acid (VPA) pharmacodynamics. A fully interactive version of this pathway is available online at PharmGKB at http://www.pharmgkb.org/pathway/PA165959313. Pharmacokinetics VPA is highly protein bound (87–95%) resulting in low clearance (6–20 ml/h/kg) [6]. There are at least three routes of VPA metabolism in humans: glucuronidation, β oxidation in the mitochondria (both considered major routes accounting for 50 and 40% of dose, respectively), and cytochrome P450 (CYP)-mediated oxidation (considered a minor route, ~10%) [7-9]. Valproate glucuronide is the major urinary metabolite of VPA (~30–50%) [8]. In-vitro studies of human liver microsomes and purified recombinant proteins have reported glucuronidation of VPA by UGT1A3, UGT1A4, UGT1A6, UGT1A8, UGT1A9, UGT1A10, UGT2B7, and UGT2B15 [8,10,11]. Other studies have disputed the role of UGT2B15, suggesting that VPA inhibits UGT2B15, but is not glucuronidated by it [12]. UGT1A1 does not have activity against VPA in vitro [8,12]. VPA is a fatty acid and can be metabolized through endogenous pathways in the mitochondria (Fig. 1). It has been observed that some of the mitochondrial metabolites of VPA generated by this pathway are hepatotoxic. The current understanding of VPA bioactivation involves the entry of 4-ene-VPA into the mitochondria, formation of a 4-ene-VPA-CoA ester with the help of ACADSB, and subsequent β-oxidation to form the reactive 2,4-diene-VPA-CoA ester [13,14]. Studies have demonstrated that the β-oxidation is blocked in fluorinated derivatives of 4-ene-VPA [15], and that the fluoro derivative of 4-ene-VPA cannot form a CoA ester [16], indicating a specific role of β oxidation of 4-ene-VPA in the formation of the 2,4-diene metabolite. This putative cytotoxic metabolite (2,4-diene-VPA-S-CoA) further gets conjugated with glutathione to form thiol conjugates. These chemically reactive metabolites generated from 4-ene-VPA have the potential to deplete mitochondrial glutathione pools [13] and form conjugates with CoA [17], in turn inhibiting enzymes in the β-oxidation pathway [18,19]. Identification of N-acetylcysteine conjugates of (E)-2,4-diene-VPA in human urine demonstrated that the reactive thiol conjugates of VPA arise primarily from the biotransformation of (E)-2,4-diene VPA in humans [20]. The key CYP-mediated branch of the VPA pathway is the generation of the metabolite 4-ene-VPA by CYP2C9, CYP2A6, and to a lesser extent by CYP2B6 [21,22]. In addition, these metabolizing enzymes also mediate the metabolism of VPA to the inactive 4-OH-VPA and 5-OH-VPA [23]. CYP2A6 also contributes partially to the formation of 3-OH-VPA [22].

A Chinese herb formula decreases the monocarboxylate transporter-mediated absorption of valproic acid in rats

Huang-Qin-Tang (HQT), a Chinese medicine prescription containing Scutellariae Radix (SR), Paeoniae Radix (PR), Glycyrrhizae Radix (GR) and JuJubae Fructus (JF), was used for the treatments of cold with symptoms of abdominalgia and diarrhea. Valproic acid (VPA) is an antiepileptic drug with narrow therapeutic window. This study investigated the effect of coadministration of HQT on the pharmacokinetics of VPA, a probe drug for monocarboxylate transporter (MCT). Rats were administered VPA alone (200.0mg/kg) and coadministered HQT (8.0g/kg) at 0.5h before VPA and 1.5h after VPA in crossover designs. In addition, the chronic effect of HQT was also evaluated by coadministration of the 7th dose at 0.5h before VPA. The serum concentration of VPA was determined by a fluorescence polarization immunoassay. The results showed that coadministration of HQT at 0.5h before VPA significantly decreased the AUC0–t and Cmax by 62% and 77%, respectively, whereas coadministration of HQT at 1.5h after VPA exerted no significant influence. When the 7th dose of HQT was given at 0.5h before VPA, the AUC0–t and Cmax of VPA were markedly decreased by 65% and 82%, respectively. Mechanism study revealed that the MCT-mediated uptake of fluorescein was inhibited by HQT and each component herbs. In conclusion, the MCT-mediated absorption of VPA was significantly decreased by concomitant administration of HQT.

Inhibition of Monocarboxylate Transporter-Mediated Absorption of Valproic Acid by Gegen-Qinlian-Tang

Valproic acid (VPA), an anti-epileptic drug with a narrow therapeutic index, is a substrate of the monocarboxylate transporter (MCT). In this study, we investigated the effect of Gegen-Qinlian-Tang (GQT), a Chinese Medicine prescription containing Puerariae Radix (PR), Scutellariae Radix (SR), Coptidis Rhizoma (CR) and Glycyrrhizae Radix (GR), on the pharmacokinetics of VPA, as a probe drug of MCT, in rats and the underlying mechanism. Sprague-Dawley rats were orally administered VPA with and without GQT in crossover design. The serum concentrations of VPA were determined by a fluorescence polarization immunoassay. The results showed that coadministration with 2.0 and 4.0 g/kg of GQT remarkably decreased the Cmax of VPA by 72% and 74% and reduced the AUC 0-t by 63% and 53%, respectively. The mechanism study using Caco-2 cells revealed that the uptake function of MCT was inhibited by GQT and each component herb. In conclusion, the MCT-mediated absorption of VPA was significantly decreased by GQT and its component herbs.

No Pharmacokinetic Interaction Between Lacosamide and Valproic Acid in Healthy Volunteers

Two open-label, randomized, multiple-dose clinical studies evaluated the potential for pharmacokinetic interaction between the antiepileptic drugs lacosamide and valproic acid. The influence of lacosamide on valproic acid pharmacokinetics (trial A) and valproic acid on lacosamide pharmacokinetics (trial B) was investigated in 32 healthy male volunteers, 16 in each trial. Volunteers in trial A received valproic acid (300 mg bid) with randomization to either early or late addition of lacosamide (200 mg bid). Those in trial B received lacosamide (200 mg bid) with randomization to either early or late addition of valproic acid (300 mg bid). Area under the concentration-time curve during a 12-hour dosing interval at steady state (AUC(τ,ss)) and maximum steady-state plasma drug concentration (C(max,ss)) were measured for each drug alone and together and tested for equivalence. The point estimates (90% confidence intervals) for AUC(τ,ss) and C(max,ss) were 104% (99%-109%) and 101% (97%-107%), respectively, for valproic acid and 100% (98%-103%) and 101% (96%-107%), respectively, for lacosamide, which were within the generally accepted equivalence range of 80% to 125%. No changes in the rate or extent of absorption, terminal half-life, or time to maximum concentration were observed. These results suggest that lacosamide and valproic acid have no relevant pharmacokinetic drug-drug interaction.

Importance of the Pharmacokinetics of Valproic Acid in an Individualized Approach to the Treatment of Epileptic Women of Fertile Age

A clinical case of the development of therapeutic side effects in an epileptic woman of childbearing age is presented. The development of therapeutic side effects occurred on the background of an intermediate therapeutic dose of a valproic acid formulation and was associated with primary (idiopathic) and secondary (valproate-induced) impairments to the folate cycle on the background of a combination of a polymorphism in the CYP2C9*3 gene and a mutation in the MTHFR gene.

Influence of UDP-glucuronosyltransferase polymorphisms on valproic acid pharmacokinetics in Chinese epilepsy patients

The aim of this study was to investigate the genetic polymorphisms of UGT1A3, UGT1A6, and UGT2B7 in Chinese epilepsy patients and their potential influence on the pharmacokinetics of valproic acid (VPA). The genetic architectures of UGT1A3, UGT1A6, and UGT2B7 in 242 epilepsy patients were detected by DNA sequencing and PCR-restriction fragment length polymorphism. Steady-state plasma concentrations of VPA in 225 patients who had received VPA (approx. 250-1,000mg/day) for at least 2 weeks were determined and associated with UGT polymorphisms. The allelic distribution of UGT1A3 in our Chinese epilepsy patients was significantly different from that in healthy subjects based on reference data. The standardized trough plasma concentration (C(S)) of VPA was much lower in our patients with the UGT1A3*5 variant than in the wild type carriers (3.24 ± 1.05 vs. 4.68 ± 1.24μg·kg·mL(-1)·mg(-1), P < 0.01). UGT polymorphisms had no influence on the pharmacokinetic interactions between carbamazepine and VPA. Our results suggest that UGT1A3*5 may be an important determinant of individual variability in the pharmacokinetics of VPA and that it may be necessary to increase VPA dose for UGT1A3*5 carriers to ensure its therapeutic range of 50-100μg/mL.

Comparison of Valproic acid Clearance between Epileptic Patients and Patients with Acute Mania.

The purpose of this study was assessment of the influence of acute manic phase on the steady state pharmacokinetics of valproic acid (VPA) in bipolar patients in comparison with those of epileptic patients. Ninteen acutely manic and 25 epileptic patients who fulfilled inclusion and exclusion criteria were entered in this prospective study. Blood samples were collected at trough time in steady state and plasma concentrations were determined by fluorescence polarization immunoassay (FPIA). VPA apparent oral clearance (CL/F) values were calculated in each patient and were compared between groups. As VPA clearance is affected by different factors such as age, total body weight, VPA dosage and the use of concurrent medications, all of these confounding factors were made similar in both groups. Comparison between two groups showed that CL/F values in acutely manic patients were significantly higher than epileptic patients (10.35±5.77 vs. 7.70±2.63 ml/kg/h, P= 0.047). Acutely manic patients require more VPA dosage to achieve serum concentrations in comparison with those found in epileptic patients. It may be suggested that this increased VPA clearance in acute manic phase may be related to abnormalities in membrane transport systems that may affect on cellular uptake of the drug and its volume of distribution. Since our study is a preliminary investigation in this field, further detailed pharmacokinetic study in acute manic patients are warranted to confirm results of this study.

A nonlinear two compartmental fractional derivative model

This study presents a new nonlinear two compartmental model and its application to the evaluation of valproic acid (VPA) pharmacokinetics in human volunteers after oral administration. We have used literature VPA concentrations. In the model, the integer order derivatives are replaced by derivatives of real order often called fractional order derivatives. Physically that means that the history (memory) of a biological process, realized as a transfer from one compartment to another, is taken into account with the mass balance conservation observed. Our contribution is the analysis of a specific nonlinear two compartmental model with the application in evaluation of VPA pharmacokinetics. The agreement of the values predicted by the proposed model with the values obtained through experiments is shown to be good. Thus, pharmacokinetics of VPA after oral application can be described well by a nonlinear two compartmental model with fractional derivatives of the same order proposed here. Parameters in the model are determined by the least-squares method and the particle swarm optimization (PSO) numerical procedure is used. The results show that the nonlinear fractional order two compartmental model for VPA pharmacokinetics is superior in comparison to the classical (integer order) linear two compartmental model and to the linear fractional order two compartmental model.

Pharmacokinetics of Valproic Acid in Guinea-pigs with Biliary Abnormality

To explore whether biliary cannulation, biliary obstruction or gall bladder obstruction could alter the disposition of valproic acid, guinea-pigs were subjected to common bile duct cannulation or ligation, gall bladder-neck ligation or sham surgery as a control. They were then given an intravenous (i.v.) dose of sodium valproate (50 mg kg-1) and the pharmacokinetics of valproic acid in each group were compared. In the cannulated group, significant decreases (P less than 0.05) in the area under the elimination curve (AUC), the volume of distribution at steady-state (Vdss) and the mean residence time (MRT) were observed. Significant increases (P less than 0.05) in the elimination rate constant (kz) and total clearance (CLtot) of valproic acid were noted. In the biliary obstructed guinea-pigs, the Vdss was significantly decreased (P less than 0.05). In the gall bladder obstructed guinea-pigs, there was a secondary peak of valproate plasma concentration, and the kz was significantly decreased. The biliary excretion of unchanged and conjugated valproic acid was 2.0 +/- 0.7 (s.e.m.) and 19.7 +/- 3.6 (s.e.m.)% of dose, respectively, and was almost completely reabsorbed in the enterohepatic recycling. Urinary excretion of unchanged and conjugated valproic acid, as well as non-conjugate metabolic clearance of valproic acid, were not significantly different among the four groups. The results suggest that the pharmacokinetics of valproic acid in guinea-pigs are particularly sensitive to interruption of the enterohepatic cycle. Biliary obstruction may elevate plasma concentrations owing to the decreased Vdss of valproic acid. Gall bladder obstruction may cause fluctuation of valproate plasma concentrations. The data indicate that the apparent total clearance of valproic acid is significantly less than the intrinsic clearance owing to enterohepatic recycling.

Phase 1 Study of Valproic Acid in Pediatric Patients with Refractory Solid or CNS Tumors: A Children's Oncology Group Report

The primary purpose of this trial was to define and describe the toxicities of oral valproic acid (VPA) at doses required to maintain trough concentrations of 100 to 150 mcg/mL or 150 to 200 mcg/mL in children with refractory solid or central nervous system (CNS) tumors. Secondary objectives included assessment of free and total VPA pharmacokinetics (PKs) and histone acetylation in peripheral blood mononuclear cells (PBMC) at steady state. Oral VPA, initially administered twice daily and subsequently three times daily, was continued without interruption to maintain trough concentrations of 100 to 150 mcg/mL. First-dose and steady-state PKs were studied. Histone H3 and H4 acetylation in PBMCs was evaluated using an ELISA technique. Twenty-six children, sixteen of whom were evaluable for toxicity, were enrolled. Dose-limiting somnolence and intratumoral hemorrhage were associated with VPA troughs of 100 to 150 mcg/mL. Therefore, the final cohort of six children received VPA to maintain troughs of 75 to 100 mcg/mL and did not experience any dose-limiting toxicity. First-dose and steady-state VPA PK parameters were similar to values previously reported in children with seizures. Increased PBMC histone acetylation was documented in 50% of patients studied. One confirmed partial response (glioblastoma multiforme) and one minor response (brainstem glioma) were observed. VPA administered three times daily to maintain trough concentrations of 75 to 100 mcg/mL was well tolerated in children with refractory solid or CNS tumors. Histone hyperacetylation in PBMCs was observed in half of the patients at steady state. Future trials combining VPA with chemotherapy and/or radiation therapy should be considered, especially for CNS tumors.

Real-time, in vivo monitoring and pharmacokinetics of valproic acid via a novel biomarker in exhaled breath

Extractive electrospray ionization mass spectrometry is shown to allow real-time, in vivo drug monitoring and pharmacokinetic measurement in a non-invasive, pain-free manner as demonstrated by the mass spectral measurement of a novel exhaled breath biomarker for valproic acid, a medication used to control epilepsy.

Simultaneous Determination of Valproic Acid and its Toxic Metabolites, 4-ene-VPA and 2,4-diene-VPA in Rat Plasma using a Gas Chromatographic-mass Spectrometric Method

− A gas chromatographic-mass spectrometric (GC-MS) method was developed for the simultaneous determination of valproic acid (VPA) and its toxic metabolites, 4-ene-VPA and 2,4-diene-VPA in rat plasma. Extraction was performed in weak acidic condition (pH 5.2) to avoid degradation of 4-ene-VPA and 2,4-diene-VPA. The recoveries for 4-eneVPA and 2,4-diene-VPA were more than 70% and that for VPA was 33-42%. R value for each compounds exceeded 0.998 in calibration curve during all the analysis. Accuracy and precision ranged from 88.3 to 113.2% and from 2.16 to 14.2%, respectively The method was successfully applied to monitor plasma concentrations of VPA, 4-ene-VPA and 2,4-diene-VPA after intravenous administration of VPA at the dose of 100 mg/kg, suggesting that these toxic metabolites may involved in the hepatotoxicity induced by VPA. Key words− valproic acid (VPA), 4-ene-VPA, 2,4-diene-VPA, quantitative analysis, GC-MS (gas chromatographic-mass spectrometry), pharmacokinetics Valproic acid (VPA, 2-propylpentanoic acid, Fig. 1) is a broad-spectrum antiepileptic agent that has a branched-chain carboxylic acid structure. VPA has been used to treat bipolar disorder, migraine, and neuropathic pain (Loscher et al., 1993; Murakami et al., 1992; Sugimoto et al., 1983), but it has been demonstrated to be associated with fatal liver toxicity (Perucca, 2002; Zimmerman et al., 1982; Zafrani et al., 1982; Bryant et al., 1986). Although the mechanism by which VPA leads to liver toxicity has not been fully elucidated, a large body of evidence suggests that reactive VPA metabolites (4ene-VPA; 2-propyl-4-pentanoic acid and its subsequent metabolite, 2,4-diene-VPA; 2-propyl-2,4-pentanoic acid) may mediate the hepatotoxicity by inhibiting mitochondrial fatty acid β-oxidation (Zimmerman et al., 1982; Gerber et al., 1979; Kassahun et al., 1991). Therefore it is important to monitor the plasma concentration of 4-ene-VPA and 2,4diene-VPA as well as VPA itself to evaluate effect and toxicity of VPA. In order to monitor plasma concentration of VPA and its toxic metabolites, it is necessary to have reliable and sensitive method for the simultaneous quantitative determination of those compounds. Because of structural similarity between fatty acid (non polar compound) and VPA, most of analytical method for VPA and/or its metabolite has been used GC-MS (gas chromatography-mass spectrometry) (Fisher et al., 1992; Yu et al., 1995; Leis et al., 2003). But those reports did not focus on the toxic metabolites, 4-ene-VPA and 2,4-diene-VPA, so it is very hard to evaluate pharmacokinetics of VPA and its toxic metabolites. Recently the method using HPLC-MS/MS (high performance liquid chromatography-tandem mass spectrometry) was reported to analyze VPA, however this method was developed only for the quantification of VPA and 4-eneVPA. Thus it is also not adequate to monitor plasma concentrations of VPA and its toxic metabolites (Cheng et al., 2007). Moreover, sample extraction with organic solvent was performed in the pH condition less than pH 3, which was not favorable condition for 4-ene-VPA and 2,4-diene-VPA since they could be degraded in acidic condition. In this study we developed adequate method for the quantitative analysis of VPA, 4-ene-VPA and 2,4-diene-VPA in rat plasma. We used mild pH condition to prevent degradation of the metabolites. This analytical method was validated and applied to evaluate pharmacokinetics of VPA and its metabolites after intravenous administration to rats. Corresponding Author : Tel : +82-2-958-5062, E-mail : jbhluck@kist.re.kr DOI : 10.4333/KPS.2010.40.3.155 156 Min Sun Lee, Young-Joo Lee, Bong Chul Chung and Byung Hwa Jung J. Pharm. Invest., Vol. 40, No. 3 (2010) Experimental

Effects of a dopamine receptor agonist and atropine sulfate on absorption of valproic acid in rats

The aim of this study is to determine whether pramipexole hydrochloride hydrate (PHH) and atropine sulfate affect valproic acid (VPA) pharmacokinetics and to evaluate how plasma VPA concentrations are altered by different PHH administration routes. The following studies were conducted on rats: 1) changes in plasma VPA concentration after simultaneous oral administration (PO) of PHH and VPA-Na; 2) effects of intraperitoneal administration (IP) of PHH on plasma VPA concentration after VPA-Na PO; 3) effects of PHH PO on plasma VPA concentration after intravenous administration (IV) of VPA-Na; and 4) changes in plasma VPA concentration after simultaneous PO of atropine sulfate and VPA-Na. Atropine sulfate PO significantly decreased the area under the concentration-time curve up to 3 h (AUC0-3, the total amount of drug plasma concentration) of VPA, suggesting that atropine sulfate decreases VPA-Na absorption probably due to reduced gastrointestinal motility by its anticholinergic action. Similarly, by PHH PO or IP, VPA AUC0-3 was significantly decreased. However, in cases of VPA-Na IV, all VPA parameters were unchanged by PHH PO. These results indicate that the PHH inhibitory effect may be caused in the absorption phase of VPA by pharmacological action of PHH, and thus PHH decreases VPA-Na bioavailability.

Plasma and cerebrospinal fluid pharmacokinetics of valproic acid after oral administration in non-human primates

Valproic acid (VPA), a widely used antiepileptic, also inhibits histone deacetylase (HDAC), and is undergoing evaluation as an anti-cancer agent. We studied the pharmacokinetics of VPA in the plasma and cerebrospinal fluid (CSF) in a non-human primate model that is highly predictive of human CSF penetration to determine if levels of VPA required to inhibit HDAC in in vivo models can be attained. Oral VPA, 75 mg/kg, was administered to four non-human primates. Serial samples of blood (n = 4) and CSF (n = 3) were obtained for pharmacokinetic studies of total and free VPA. Plasma and CSF VPA concentrations were measured using the commercially available Abbott AxSYM VPA assay reagent system (Abbott Laboratories, Abbott Park, IL, USA). The resultant plasma and CSF data were evaluated using pharmacokinetic modeling methods. At a dose of 75 mg/kg, the maximum plasma concentration of VPA was 130.1 +/- 70.6 microg/ml (mean +/- standard deviation) for total drug and 53.3 +/- 44.4 microg/ml for free drug. The mean plasma area under the curve (AUC) for total drug was 680 +/- 233 microg/ml h and for free drug 146 +/- 89 microg/ml hr. The maximum CSF concentration occurred 2-3 h after administration and was 28.2 +/- 18.6 microg/ml. The CSF AUC for VPA was 108 +/- 52 microg/ml h. The CSF penetration of VPA was 12.9 +/- 5.1% for total drug and 57.0 +/- 8.7% for free drug. Disappearance from the plasma followed non-linear kinetics with a V (max) of 321.2 +/- 65.6 microg/kg/min and a K (m) of 17.2 +/- 13.7 mg/l. Valproic acid deserves further study for the treatment of CNS tumors given its high CSF penetration after oral dosing coupled with the anti-tumor activity observed in preclinical studies.

Effects of Tandospirone Citrate and Oxazolam on Pharmacokinetics of Valproic Acid Following Oral Administration of Sodium Valproate to Rats

The drug interactions between tandospirone citrate,a non-benzodiazepine antianxiety drug,and oxazolam,a minor benzodiazepine tranquilizer and antianxiety agent,and valpronic acid (VPA),an anti-convulsant,were studied in rats.When tandospirone citrate or citric acid was administered orally immediately following oral administration (p.o.) of sodium valproate (VPA-Na),the plasma VPA levels were significantly lower than those in the control.In addition,pharmacokinetic parameters such as plasma VPA concentration and area under the plasma concentration-time curve up to 3 hr (AUC) were significantly decreased.However,when tandospirone citrate was administered intraperitonally,or both it and oxazolam were administered orally with VPA-Na,the VPA pharmacokinetic parameters were unchanged.These results suggest that decreases in plasma VPA concentrations (including those for AUC) may be due to a reduction in VPA-Na adsorption from the intestinal tract due to tandospirone citrate.