In vivo activity and hydrophobicity of cytostatic aziridinyl quinones.

For a series of 3,6-disubstituted bisaziridinylbenzoquinones the in vivo and in vitro activities against murine tumors, as well as the in vivo toxicity, are analyzed. Properties describing biochemical and physicochemical reactions are also incorporated in the analyses. The important 1-octanol/water partition coefficients were determined, using a fast variation of the shake flask method. New pi'-values were calculated for the substituents in this series. These quinone pi'-values deviate strongly from the standard pi-values, especially for hydrogen-bonding substituents. To discriminate between the toxic and therapeutic activity of the compounds, principal components and partial least squares analyses were applied. Evidence is presented for selective antitumor action of the investigated compounds. The L1210 clonogenic assay only seems to relate to the general cytotoxicity and has no predictive value for in vivo activity for these compounds. The activity is correlated to the hydrophobicity of the quinones. The toxicity correlates with the ease of reduction, contrary to the hypothesis of bioreductive activation as a mechanism for selectivity.

Liquid chromatographic analysis of carboxylic acids using N-(4-aminobutyl)-N-ethylisoluminol as chemiluminescent label: determination of ibuprofen in saliva.

N-(4-Aminobutyl)-N-ethylisoluminol was used for labelling of carboxylic acids. The derivatization reaction was carried out with 1-hydroxybenzotriazole as pre-activator of the carboxylic acid function and N-ethyl-N'-(3-dimethylaminopropyl)carbodiimide as the coupling reagent. Optimum conditions for the derivatization were determined by using factorial design analysis, with ibuprofen as the test compound. Chemiluminescence detection was carried out using a post-column on-line electrochemical hydrogen peroxide generation system and the addition of microperoxidase as the catalyst. The detection limit of derivatized ibuprofen in human saliva was 0.7 ng per 0.5 ml of saliva, with a recovery of 96.1 +/- 1.3%. The method was linear over at least three decades (2.5 ng to 2.5 micrograms) and the repeatability was satisfactory (R.S.D. = 5.2% at the 25 ng level; n = 4).

Absolute bioavailability and pharmacokinetics of oral teniposide.

The absolute bioavailability and pharmacokinetics of orally administered teniposide were investigated in 25 patients. All patients received 50 to 60 mg/m2 teniposide intravenously on day 1, before oral administration. Six patients received 60 mg/m2 as a single oral dose on day 8; 5 patients received 60 mg/m2 and 120 mg/m2 as a single oral dose on days 8 and 15, respectively; 5 patients received 120 mg/m2 and 240 mg/m2 as a single oral dose on days 8 and 15, respectively; 6 patients received 60 mg/m2 as a single oral dose on 5 consecutive days from days 8 to 12; and 3 patients received 50 mg/m2 three times a day at 6-hour intervals on day 8. The mean absolute bioavailability was 41.6% +/- 14.2% with a large interindividual variability (range, 19.7% to 71.4%) and a low intraindividual variability (range, 2.8% to 13.9%). At a dose of 240 mg/m2, the bioavailability was decreased, whereas administration of multiple doses on 1 day or 5 consecutive days increased the overall bioavailability. In conclusion, teniposide can be administered orally with a bioavailability comparable with that of etoposide. The schedule dependency of both drugs warrants investigations of oral administration for 21 or more days. A formulation of teniposide capsules of 50 mg or less would be most helpful to facilitate oral administration.

Comparative study on the determination of the anti-neoplastic drug teniposide in plasma using micellar liquid chromatography and surfactant-mediated plasma clean-up.

The potential of micellar liquid chromatography and of an on-line surfactant-mediated sample cleanup, which involves column-switching prior to conventional reversed-phase high-performance liquid chromatography, has been evaluated for the determination of the anti-neoplastic drug teniposide in plasma by using electrochemical detection. A major advantage of surfactant-mediated techniques is that they allow fully automated processing of plasma samples, because protein precipitation is prevented by the addition of the surfactant sodium dodecylsulphate. With the automated column-switching technique, a degree of sample enrichment and of selectivity can be attained, which is similar to that for the conventional procedure which, however, involves a labour-intensive off-line isolation of teniposide, using liquid-liquid extraction prior to chromatography. An inherent drawback of automated micellar liquid chromatography is that no sample clean-up or preconcentration can be carried out, which results in only a moderate detection limit and selectivity. The linearity, reproducibility and recovery of the surfactant-mediated techniques are similar to those of the conventional procedure. Based on the presented results, it was concluded that the surfactant-mediated column-switching technique is a highly attractive sample enrichment technique with respect to simplicity, speed and cost.

Automated high-performance liquid chromatographic determination of plasma free fatty acids using on-line derivatization with 9-bromomethylacridine based on micellar phase-transfer catalysis.

The on-line use of micellar phase-transfer catalysis is described for the automated reversed-phase high-performance liquid chromatographic (RP-HPLC) determination of free fatty acids in plasma; minimum manual sample handling is involved. After diluting plasma ten-fold with the aqueous micellar system, which contains 25 mM of the non-ionic surfactant, Arkopal N-130 and 6 mM of the ion-pair agent tetrakis(decyl)ammonium bromide, the reaction of the fatty acids with the fluorophore 9-bromomethylacridine is complete within 5 min at 60 degrees C. Prior to RP-HPLC separation, interfering proteins are removed using an on-line filter and a column-switching unit. More than 100 samples can be injected onto a single pre-column. The detection limit is ca. 300 nM; the precision is better than 3% using an internal standard.

High-performance liquid chromatographic determination of amantadine in urine after micelle-mediated pre-column derivatization with 1-fluoro-2,4-dinitrobenzene.

Cationic micelles have been used for the derivatization of the anti-Parkinson drug amantadine with the chromophore 1-fluoro-2,4-dinitrobenzene in urine. In the presence of 90 mM cetyltrimethylammonium bromide (CTAB), the conversion of amantadine into its derivative is complete within 4 min at 60 degrees C and pH 11. Such a short reaction time allows a fully automated pre-column derivatization of amantadine in an on-line combination with reversed-phase high-performance liquid chromatography. This cannot be attained when using purely aqueous derivatization mixtures because then the reaction takes some 20 min at the same temperature. Without the use of an internal standard, the repeatability of the automated determination at the 0.5 microgram ml-1 level is ca. 6%, whilst the detection limit is 75 ng ml-1 (S/N = 3). The present study clearly demonstrates that micellar systems can be beneficially used for the on-line precolumn derivatization of amines in urine.

The bioavailability of three altretamine formulations.

The bioavailability of two altretamine preparations was studied in a randomized cross-over design. The two preparations were compared with a third in a parallel design. Dissolution differences between the preparations were observed, which could give rise to differences in bioavailability caused by the extensive first-pass effect of altretamine. The in vivo data showed a trend to differences in bioavailability.

Automated reversed-phase chromatographic analysis of etoposide and teniposide in plasma by using on-line surfactant-mediated sample clean-up and column-switching.

An automated high-performance liquid chromatographic method for the plasma assay of two neutral drugs, etoposide and teniposide, involving direct plasma injection is presented. The problematic nature of protein precipitation has been circumvented by adding the anionic surfactant sodium dodecyl sulphate to the plasma at a final concentration of 38 mM. Plasma samples are loaded on to a clean-up column with an aqueous mobile phase with which the analyte(s) is (are) retained, whereas the solubilized plasma proteins are flushed to waste. Next, the retained compounds are eluted from the clean-up column on to the analytical column by using the chromatographic mobile phase with a higher elution capacity. The column-switching technique is used to achieve an automated assay. At least 10 ml of plasma, representing 100 repeated injections of 100 microliters or five repeated injections of 2 ml, can pass through the clean-up column without increasing the back-pressure. The recovery increased considerably from 10-30% to 90-95% on adding surfactant to the plasma samples prior to the analysis. The relative standard deviation of the proposed clean-up procedure is 3.5% (n = 6) for both drugs measured at the 2 micrograms/ml level without using an internal standard. The limit of determination with 100-microliters injections is 0.10-0.15 microgram/ml for ultraviolet detection and is seven times lower with electrochemical detection. Teniposide was determined in patients' plasma and the results agreed well with those obtained by the conventional procedure involving manual liquid-liquid extraction prior to chromatographic analysis.

High-performance liquid chromatographic determination of valproic acid in plasma using a micelle-mediated pre-column derivatization.

A method for the determination of valproic acid (2-propylpentanoic acid) in plasma by high-performance liquid chromatography (HPLC) after pre-column derivatization is described. The derivatization of valproic acid with a fluorophore and UV label, 4-bromomethyl-7-methoxycoumarin, is performed in plasma diluted with an aqueous micellar system. No extraction or solvent evaporation steps are required. The mechanism of the derivatization of the carboxylic acid is based on phase-transfer catalysis. The sample preparation, including the derivatization step, is rapid and very simple. The proposed HPLC-method was evaluated and compared with a standard immunological assay used for the determination of valproic acid in plasma.

Determination of vinca alkaloids in plasma and urine using ion-exchange chromatography on silica gel and fluorescence detection.

The development of a method for the determination of the antineoplastic vinca alkaloids vinblastine and vindesine in biological samples is described. The selectivity of the assay is high owing to the use of solid-phase extraction on a cyanopropyl extraction column prior to isocratic chromatography on unmodified silica gel with fluorescence detection. The influence of acetonitrile concentration and mobile phase pH on the capacity factors of the drugs was studied in order to optimize the separation between the drugs and endogenous components. The effect of varying the type and concentration of competing cations in the mobile phase was also examined. The limit of determination (signal-to-noise ratio = 3) for vinblastine is 0.5 ng/ml in plasma and urine and for vindesine 2.5 ng/ml. The assay is suitable for determining the concentrations of both compounds in plasma and urine samples from patients.

Determination of delta 9-tetrahydrocannabinol in plasma using solid-phase extraction and high-performance liquid chromatography with electrochemical detection.

A method for the determination of nanogram amounts of delta 9-tetrahydrocannabinol (THC) in plasma and serum is described. THC was quantitatively isolated by solid-phase extraction after addition of an aqueous solution of urea and methanol to the sample. The extracts were analysed by high-performance liquid chromatography with electrochemical detection in the oxidizing mode. The detection limit of THC is ca. 100 pg for a signal-to-noise ratio of 3. With this method, levels of 2 ng/ml of THC in plasma can be measured.

Fractionated total body irradiation and high-dose VP 16-213 followed by allogeneic bone marrow transplantation in advanced leukemias.

Thirty-eight patients (median age, 21 years) with acute nonlymphoblastic leukemia (ANLL) (17 patients), acute lymphoblastic leukemia/lymphoma (ALL) (18 patients), chronic myelogenous leukemia (two patients), and refractory anemia received allogeneic bone marrow transplants from HLA-identical sibling donors or a one-antigen-mismatched brother (one patient) after a preparatory regimen consisting of fractionated total body irradiation and high-dose VP 16-213 (60 to 70 mg/kg body weight). Of the 33 patients with acute leukemia who received grafts from HLA-identical donors, three patients with ANLL received transplants in first remission and one patient with standard-risk ALL received a graft while in second remission. All other patients were in more advanced stages of their disease or exhibited other high-risk features. At the time of analysis, 20 of the 33 patients were alive, with 19 of them remaining in continued complete remission for 6 to 35 months (median, 18 months). The 3-year actuarial disease-free survival rate of 56.6% +/- 9.7% (SE) and the actuarial relapse rate of 11.9% +/- 6.8% (SE) demonstrate that the combination of fractionated total body irradiation and high-dose VP 16 is an effective mode of therapy in patients with advanced leukemias. Preliminary experience cautions against the use of VP 16 instead of cyclophosphamide in any clinical situation carrying an increased risk of graft rejection because the immunosuppressive potency of VP 16 is largely untested but may be inferior to that of cyclophosphamide.

Etoposide and teniposide. Bioanalysis, metabolism and clinical pharmacokinetics.

Etoposide (VP 16-213) and teniposide (VM 26) are semisynthetic epipodophyllotoxin derivatives active against a variety of tumours. The clinical efficacy has led to an increasing interest in these compounds. This review presents information on the mechanism of action, biochemical pharmacology, bioanalysis, metabolism and pharmacokinetics of etoposide and teniposide.

Study of the derivatization of n-alkylamines with 1-fluoro-2,4-dinitrobenzene in the presence of aqueous cetyltrimethylammonium bromide micelles.

The use of aqueous cetyltrimethylammonium bromide micelles in the derivatization of n-alkylamines with 1-fluoro-2,4-dinitrobenzene was investigated systematically. The rate constants of derivatization of the n-alkylamines (C1-C8) were analysed using liquid chromatography. Up to butylamine the micellar rate enhancement depends on the electrostatic interactions between the amines and cetyltrimethylammonium bromide, and beyond C4 it depends mainly on the hydrophobic interactions. The reaction rates are also enhanced by a micelle-induced decrease of the pKa of the amines, but to a lesser extent. The derivatization rates for the longer alkylamines are comparable with those in dipolar aprotic solvents. Pharmaceutical and biomedical science is likely to benefit from the use of micellar systems in pre-column derivatization reactions in aqueous solutions.

Analysis of vinca alkaloids in plasma and urine using high-performance liquid chromatography with electrochemical detection.

The reversed-phase high-performance liquid chromatography with electrochemical detection was used to quantify plasma and urine levels of vinblastine, vincristine, vindesine and a metabolite of vinblastine, desacetylvinblastine. Sample clean-up consisted of solid-phase extraction with a Bond Elut CN column. The extracts were separated on a Hypersil ODS column. The mobile phase consisted of a mixture of methanol and 10 mM phosphate buffer (pH 7.0). The limit of sensitivity using electrochemical detection was 100 pg on-column for all compounds with a signal-to-noise ratio of 3. Quantification of the compounds in human plasma and urine was possible down to 1 ng/ml (ca. 1 pmol). Pharmacokinetic results show that the sensitivity of the method is adequate for drug monitoring in clinical research.

Pharmacokinetics of high-dose teniposide.

This paper describes the pharmacokinetics of teniposide (VM-26) after being administered iv in high doses to eight cancer patients (maximum dose, 1.0 g/m2). VM-26 levels in plasma, urine, saliva, duodenal fluid, and cerebrospinal fluid were determined using high-performance liquid chromatography in combination with electrochemical detection. The plasma concentration-time curve of VM-26 showed a triphasic decay with a slow third phase in five patients, whereas in two patients the plasma concentration decay was biphasic. The plasma pharmacokinetics of VM-26 proved to be linear and could be fitted to a three-compartment model (five patients) and to a two-compartment model (two). The steady-state volume of distribution varied from 13.2 to 24.7 L/m2. The total-body clearance ranged from 5.84 to 10.18 ml/minute/m2. Low concentrations of VM-26 were found in saliva, duodenal fluid, cerebrospinal fluid, and urine. Excretion of unchanged VM-26 into the urine varied from 8.8% to 13.9% of the administered dose. No glucuronide of VM-26 could be detected in plasma or other biological fluid.