Cork-based activated carbons as supported adsorbent materials for trace level analysis of ibuprofen and clofibric acid in environmental and biological matrices.

In this contribution, powdered activated carbons (ACs) from cork waste were supported for bar adsorptive micro-extraction (BAµE), as novel adsorbent phases for the analysis of polar compounds. By combining this approach with liquid desorption followed by high performance liquid chromatography with diode array detection (BAµE(AC)-LD/HPLC-DAD), good analytical performance was achieved using clofibric acid (CLOF) and ibuprofen (IBU) model compounds in environmental and biological matrices. Assays performed on 30 mL water samples spiked at the 25.0 µg L(-1) level yielded recoveries around 80% for CLOF and 95% for IBU, under optimized experimental conditions. The ACs textural and surface chemistry properties were correlated with the results obtained. The analytical performance showed good precision (<15%), suitable detection limits (0.24 and 0.78 µg L(-1) for CLOF and IBU, respectively) and good linear dynamic ranges (r(2)>0.9922) from 1.0 to 600.0 µg L(-1). By using the standard addition methodology, the application of the present approach to environmental water and urine matrices allowed remarkable performance at the trace level. The proposed methodology proved to be a viable alternative for acidic pharmaceuticals analysis, showing to be easy to implement, reliable, sensitive and requiring low sample volume to monitor these priority compounds in environmental and biological matrices.

[Conjugation reactions of ciprofibrate with human and laboratory animals]. / Konjugationsreaktionen von Ciprofibrat bei Menschen und Versuchstieren.

An open problem of the lipid lowering agent ciprofibrate (rac-2-[4-(2,2-dichlorocyclopropyl)-phenoxy]-2-methylpropanoic acid, CAS 52214-84-3) is its metabolism concerning the conjugation with amino acids and glucuronic acid. It could be solved by syntheses of the needed reference compounds--unknown up to now--and administration of ciprofibrate to volunteers and rats. Unexpectedly the conjugation compounds with amino acids are stable in vitro and in metabolism. There was no evidence for any conjugation reaction with amino acids by investigating samples of urine and faeces. On the contrary the urine of humans contains 90-97% of beta-O-acylglucuronide, whereas rat urine shows only 10% of the calculated amount.

Achiral and chiral determination of ciprofibrate and its glucuronide in human urine by capillary electrophoresis.

A method for achiral separation of the racemic hypolipidaemic agent ciprofibrate and its main metabolite ciprofibrate glucuronide by capillary electrophoresis (CE) was developed. The glucuronide was isolated from urine by chromatographic procedures and characterized by alkaline as well as enzymatic hydrolysis and mass spectrometric and nuclear magnetic resonance experiments. Chiral discrimination of the ciprofibrate enantiomers and their diastereomeric glucuronides by CE was achieved by the use of gamma-cyclodextrin as buffer additive. The fractionated crystallization of ciprofibrate yielded the R-(+)-enantiomer as less soluble diastereomeric salt with (+)-1-phenylethylamine. This allowed the identification of the enantiomers of ciprofibrate as well as the diastereomeric glucuronides of ciprofibrate by CE. In a study with three volunteers inter- and intra-individual differences of ratios of both ciprofibrate glucuronides in urine were observed. After oral administration of a single dose of the racemate the S-ciprofibrate glucuronide was mainly excreted in the first time intervals, in the last time intervals the R-glucuronide dominated.

[The pharmacokinetics of hypolipemic agents. 10. The dehalogenation of the hypolipemic agent ciprofibrate]. / Zur Pharmakokinetik von Lipidsenkern, 10. Mitt.: Zweiter Beitrag zur Frage der Dehalogenierung des Lipidsenkers Ciprofibrat.

rac-2 described as a metabolite of rac-1 was synthesized in four steps starting with rac-3. Partial dehalogenation occurs with LiAlH4. A new structure assignment of the resulting stereoisomers resulted from NMR spectroscopy. After oral administration of rac-1 in multiple dose studies to volunteers, rac-2 could not be detected within the limitations of sensitivity of HPLC (UV-detector) in plasma or in urine.

Further studies on the involvement of selenium in peroxisome proliferation in rat liver. Comparison of effects with clofibric acid and perfluorooctanoic acid and the pharmacokinetics of [14C]clofibrate.

Most effects of the peroxisome proliferator clofibrate on rat liver are marginal or absent in selenium (Se) deficiency. The purpose of the present study was to determine whether the uptake or distribution of clofibrate is altered by Se deficiency. Rats were fed a Se-adequate or -deficient diet for 10-11 weeks and then these same diets with 0.5% (w/w) clofibric acid (the direct acting hydrolysis product of clofibrate) or 0.02% (w/w) perfluorooctanoic acid (PFOA) for 10 days. Other groups of rats received radiolabeled clofibrate by intubation. Clofibric acid was an ineffective as clofibrate in producing effects (i.e. decreased body weight gain, increases in liver somatic index and protein content of the mitochondrial fraction, and increased activities of catalase and peroxisomal fatty acid beta-oxidation) in the liver of Se-deficient rats. Microsomal omega-hydroxylation was, however, equally induced in both dietary groups. In contrast to clofibric acid, the biological effects of PFOA were not affected by Se status. Furthermore, neither the tissue distribution (plasma, liver and kidney) nor the urinary excretion of 14C was affected by Se deficiency. These results demonstrate that the hydrolysis of clofibrate to clofibric acid is not impaired in the Se-deficient rat. In addition, the involvement of Se in the effects of peroxisome proliferators differs for different members of this structurally heterogeneous group of compounds. It is concluded that the Se-deficient rat may provide valuable information concerning the biochemical mechanism(s) underlying peroxisome proliferation.

Oral contraceptives stimulate the excretion of clofibric acid glucuronide in women and female rats.

1. Glucuronidation of clofibric acid, the pharmacologically active form of the hypolipidemic drug clofibrate was investigated in a human population, either in vitro with liver homogenates from biopsies, or after ingestion of the drug and determination of the urinary metabolite. No difference in the glucuronidation rate according to age of the patients was observed. Bilirubin but not clofibric acid glucuronidation was significantly higher in women (106% increase), when expressed per gram of tissue. 2. The excretion of clofibryl glucuronide in women who took oral contraceptives was significantly enhanced by 25%. 3. In female rats, treatment with the contraceptive agent norethindrone also stimulated by 48% the formation of clofibrylglucuronide in liver microsomes.

PIP: The effect of oral contraceptive (OC) intake on the glucuronidation of clofibric acid was investigated both in vivo, by measuring the excretion of urinary clofibrylglucuronide in female Wistar rats, or in vitro, with liver biopsy homogenates from 69 health female volunteers. In both study groups norethindrone markedly enhanced glucuronidation of clofibric acid while the situation was modulated by the administration of ethinyl estradiol. Excretion of clofibric acid into urine averaged 142.1 + or - 38.4 mg in OC users compared to only 105.8 + or - 31.5 mg in non-users. When expressed per gram of tissue, bilirubin but not clofibric acid glucuronidation was significantly greater in human liver biopsies from OC users compared to non-users. In rats treated with norethindrone, the liver microsomal protein content was increased 28% over the average control value and formation of clofibylglucoride was stimulated by 48%. It is hypothesized that the stimulation of clofibylglucuronide in OC users results from the selective stimulation in the liver of the isoenzyme involved in the process of the glucuronidation of clofibric acid.

Urinary glucuronide excretion of fenofibric and clofibric acid glucuronides in man. Is it polymorphic?

The possible polymorphism of the glucuronidation reaction in man has been investigated using two hypolipidaemic compounds, fenofibrate and clofibrate, as the test probes. The formation of fenofibryl and clofibryl glucuronides was identified by their susceptibility to hydrolyses by beta-glucuronidase. The urinary excretion of the glucuronides was measured in 72 healthy volunteers after a single dose of fenofibrate, and in 104 subjects given a single dose of clofibric acid. Fenofibrate was excreted at a lower rate than clofibrate, since 13.94% and 26.55% of the doses of fenofibrate and clofibrate respectively, were recovered in urine in 8 h. Correlation analysis indicated that sex and body mass index significantly influenced the formation of fenofibryl glucuronide, whereas age and oral contraceptives affected the excretion of clofibryl acid glucuronide. The 8-hour urinary excretion patterns of clofibryl glucuronide and of clofibric acid presented a Gaussian distribution, whereas those of fenofibryl glucuronide and fenofibric acid showed 2 populations. When the metabolic ratio free fenofibric acid/glucuronide was considered, 84.7% of subjects presented the ratio 0.147, and 15.3% had the 3-fold higher ratio of 0.421. The study has shown, in the human population studied, that the glucuronidation of fenofibric acid but not that of clofibric acid may present a polymorphism.

The influence of renal insufficiency and haemodialysis on the kinetics of ciprofibrate.

1. The kinetics of the hypolipidaemic drug, ciprofibrate, were studied after a single oral dose (100 mg) in subjects with normal renal function (n = 6), patients with mild (n = 6) and severe (n = 6) renal insufficiency as well as in haemodialysed patients (n = 5). 2. Under fasting conditions, ciprofibrate, was absorbed rapidly in subjects with normal renal function, and its apparent elimination half-life was approximately 81 h. Both renal clearance (0.15 ml min-1) and cumulative renal excretion (less than 7% of the administered dose) were low. 3. Mild renal insufficiency did not alter the pharmacokinetics of ciprofibrate, but severe renal impairment significantly reduced both its renal clearance and cumulative urinary excretion and increased the apparent elimination half-life. 4. A 5 h haemodialysis session did not lower the plasma concentrations of ciprofibrate. 5. It is concluded that, from a pharmacokinetic point of view, a reduction in the dosage of ciprofibrate should be considered in patients with a glomerular filtration rate below 30 ml min-1/1.73 m2.

Evaluation of a versatile reversed-phase high-performance liquid chromatographic system using cethexonium bromide as ion-pairing reagent for the analysis of glucuronic acid conjugates.

An ion-pair high-performance liquid chromatographic technique has been developed to reduce the elution times of both glucuronic acid conjugates and their free aglycones. The chromatographic system combines the use of a reversed-phase column (LiChrospher CH-18; 5 microns) and a mobile phase of methanol (70-80%)-0.01 M phosphate buffer (pH 6.0) containing 2.5 mM cethexonium bromide as counter-ion at a flow-rate of 1 ml min-1. The hydrophobicity of this quaternary ammonium ion-pairing reagent and the high content of the organic modifier in the mobile phase provide close and short elution times for a wide structural variety of compounds (i.e. alcohols, phenols, steroids, carboxylic acids) and their conjugates with glucuronic acid (capacity factors lower than 7.5), without compromising the selectivity with respect to endogenous compounds of the microsomal incubation medium and urine. Advantages of cethexonium bromide over conventional tetrabutylammonium salts are clearly demonstrated, and the described system was applied to the simultaneous quantitation of clofibric acid and its acylglucuronide in human urine and validated for a pharmacogenetics purpose.

Influence of food on the absorption of the p-chlorophenolic ester of chlorophenoxyisobutyric acid in man.

A study was designed to investigate the effect of a fatty meal on the absorption of chlorophenoxyisobutyric acid (CPIB) in six healthy adult volunteers after oral administration of the p-chlorophenolic (PCP) ester of CPIB. Plasma concentrations of CPIB when administered with food were higher than those observed in the fasting state. The Cmax in the former case was 24.0 +/- 4.6 mg l-1 as against a value of 15.2 +/- 6.9 mg l-1 in the latter (P less than 0.01). The pharmacokinetic parameters measured were found to be linear with the dose administered. This could be due to low plasma concentrations of CPIB unable to cause a saturation of the plasma protein binding of this drug. It is concluded that a fatty meal enhances the absorption of this hypolipidaemic drug.

The analysis of [14C]clofibric acid glucuronide and [14C]clofibric acid in plasma and urine using high-performance liquid chromatography.

A high-performance liquid chromatographic method has been developed for the quantitation of [14C]clofibric acid glucuronide and [14C]clofibric acid using conventional scintillation counting. The assay has a linear relationship between the added and observed ratios of clofibric acid glucuronide: clofibric acid in the range of 0.001-0.6 for plasma and 0.5-100 for urine, and is able to quantitate previously unmeasurable concentrations of clofibric acid glucuronide in plasma.

The simultaneous analysis of clofibric acid and probenecid and the direct analysis of clofibric acid glucuronide by high-performance liquid chromatography.

A rapid and selective high-performance liquid chromatographic method for the simultaneous quantitative analysis of clofibric acid and probenecid in plasma and urine and for the direct analysis of clofibric acid glucuronide in plasma is described. Both methods involve direct injection of deproteinised body fluids. Concentrations of as low as 10 mg/l of clofibric acid and probenecid and 1.5 mg/l of clofibric acid glucuronide can be measured by the analysis. The coefficient of variance for these methods ranges from 1--7%.

Clinical pharmacokinetics of bezafibrate in patients with impaired renal function.

The pharmacokinetics of the new lipid-lowering drug bezafibrate has been investigated in patients with impaired renal function and hyperlipoproteinaemia. 12 patients received a single oral dose of bezafibrate 300 mg. Plasma and urine samples were collected and bezafibrate was analyzed by gas chromatography. Eight of the patients had moderately impaired renal function, with a creatinine clearance between 20 and 40 ml/min; the mean plasma half-life of bezafibrate in them was 7.8 +3.9 h (SD) and the plasma clearance was 0.03 +0.02 1/kg . h. Three of the patients had a creatinine clearance greater than 40 ml/min; in them the plasma half-life was shorter, 4.6 +1.2 h, and the plasma clearance was higher, 0.06 +0.01 1/kg . h. The slowest elimination of bezafibrate was found in a patient with a creatinine clearance of only 13 ml/min. This patient had a plasma half-life of 20.l h, which is ten times longer than has been reported in healthy volunteers. Thus, when treating hyperlipoproteinaemia in patients with impaired renal function, the dosage of bezafibrate must be individualized because of its reduced renal elimination.

Comparative pharmacokinetics of 400 mg bezafibrate after a single oral administration of a new slow-release preparation and the currently available commercial form.

Bezafibrate is a new lipid-lowering agent with a more pronounced pharmacological effect, a different metabolism and a much shorter apparent half-life than clofibrate. The serum concentration curves and the urinary excretion were determined in seventeen healthy volunteers after administration of 400 mg of a new slow-release preparation. The results were compared with those of a similar experiment in twenty healthy volunteers who received 400 mg as a single dose in the form of 2 tablets of Cedur, which is the currently marketed form in Germany (each tablet contains 200 mg bezafibrate). The areas under the serum concentration curves were not statistically significantly different (median of 31.3 mg . h/l for the slow-release and of 39.4 mg . h/l for the marketed preparation). Renal clearance differed only within the margin of biological variation. It can be assumed that the relative biological availability of both preparations is similar. In none of the volunteers were subjective or objective side-effects observed.