Fluorogenic derivatization of peptides with naphthalene-2,3-dicarboxaldehyde/cyanide: optimization of yield and application in the determination of leucine-enkephalin spiked human plasma samples.

Initial attempts to derivatize the alpha-amino site of several tripeptides with naphthalene-2,3-dicarboxaldehyde/cyanide (NDA/CN) resulted in poor yields of the expected N-substituted 1-cyanobenz[f]isoindole (CBI) products. Examination of the CBI-formation mechanism, in conjunction with knowledge of the general structure-reactivity properties of the tripeptides, led to the recognition of a competing non-productive reaction pathway. Through the use of model reactions and the isolation and structural elucidation of a predicted side-product the viability of the competing pathway was confirmed.

Liquid chromatographic determination of cyclophosphamide enantiomers in plasma by precolumn chiral derivatization.

On the basis of reactions described in the synthetic literature, a two-step chiral derivatization sequence was developed for the anticancer agent cyclophosphamide (CP). The sequence involves amidoalkylation of CP with anhydrous chloral containing 1% dimethylformamide followed by acylation of the resulting secondary alcohol with a chiral carboxylic acid chloride, (+)-6-methoxy-alpha-methyl-2-naphthaleneacetyl chloride, to form a diastereomeric pair. Derivatized (-)-CP and (+)-CP exhibited retention times of 17.

Analytical challenges to the pharmaceutical industry in developing products of biotechnology.

Problems associated with the analysis of peptides and proteins in pharmaceutical products produced by biotechnology are discussed. Analytical techniques for the determination of peptides and proteins in such products are classified into four categories: bioassays, binding assays, enzyme assays and physical/chemical methods.

Suitability of DTAF as a fluorescent labelling reagent for direct analysis of primary and secondary amines--spectral and chemical reactivity considerations.

DTAF has been used successfully to prepare fluorescent labelled reagents for fluorescence polarization immunoassays. Its applicability as a derivation reagent for direct fluorescence analysis of primary and secondary amines was evaluated. DTAF was shown to have spectral properties that closely resemble those of fluorescein and that are apparently insensitive to the presence of the triazine nucleus.

Analysis by liquid chromatography of desipramine in aqueous solutions and plasma application of dichlorotriazinylaminofluorescein (DTAF) as a pre-column fluorescent derivatising agent for selected secondary amines.

Dichlorotriazinylaminofluorescein (DTAF) has been proposed as a fluorescent derivatising agent for secondary aliphatic amines. In this report, its applicability as a pre-column reagent is demonstrated by its use in the analysis of aqueous solutions and plasma samples of the tricyclic antidepressant, desipramine. Samples were processed by liquid-solid extraction onto XAD-2 resin, followed by DTAF derivatisation of the fraction of the column effluent containing analyte.

Chemical derivatization as a strategy to enhance detectability of agents used in cancer management.

The bioanalysis of drugs used in the management of cancer is often complicated by the lack of selectivity and sensitivity. Chemical derivatization of these drugs prior to their chromatographic analysis represents a viable strategy to improve chromatographic resolution and to enhance detectability. This review provides examples of how this approach can meet these objectives.

The application of chemical derivatization to clinical drug analysis.

The sensitivity and selectivity achievable in the analysis of drug substances from biological matrices is often limited by the physical and chemical properties of the analyte. These limitations are further exacerbated by the inherent reactivity of most drugs in biological systems (i.e.

Design and synthesis of a theophylline bonded-phase column for HPLC--application to separation of aromatic carboxylic acids.

A stationary phase has been designed and synthesized in which theophylline residues are covalently bonded to a silica support through an eight carbon hydrocarbon linkage. The phase offers improved resolution in the separation of aromatic carboxylic acids over that available with conventional reversed phase supports. The column is relatively stable.

Aspects of the stability of isoindoles derived from the reaction of o-phthalaldehyde-ethanethiol with primary amino compounds.

The stability of a series of fluorescent isoindoles formed under analytical conditions following the reaction of o-phthalaldehyde (OPA) and ethanethiol (ET) with a series of primary amines is reported. Increasing the bulk and degree of substitution of the isoindole N-substituent resulted in substantial increases in isoindole stability. The effects of excess reagents on isoindole stability is examined and OPA is observed to accelerate isoindole degradation whilst ET provides a stabilizing effect.

Phase I evaluation and pharmacokinetics of tiazofurin (2-beta-D-ribofuranosylthiazole-4-carboxamide, NSC 286193).

Tiazofurin (2-beta-D-ribofuranosylthiazole-4-carboxamide, TCAR, Riboxamide, NSC 286193) is a novel C-nucleoside with antitumor activity against several murine tumor models, including Lewis lung carcinoma. The mechanism whereby this compound exerts its antineoplastic effects is most likely related to a state of guanine nucleotide depletion whereby the anabolite, thiazole-4-carboxamide adenine dinucleotide, potently inhibits inosine-5'-monophosphate dehydrogenase. This Phase I study was designed to determine the maximally tolerated dose of Tiazofurin administered on a 5-day, every-28-day schedule.

Rational design and evaluation of improved o-phthalaldehyde-like fluorogenic reagents.

Evidence was presented suggesting that the fluorescent isoindole produced by reaction of o-phthalaldehyde (OPA), ethanethiol, and primary amine was formed by initial imine formation followed by conversion to an alpha-alkylaminobenzylsulfide and subsequent ring closure to form the isoindole nucleus. This mechanism suggested that the minimum structural requirement for condensation to an isoindole was an o-diacyl benzene in which one of the carbonyl groups was aldehydic. A major drawback of OPA as an analytical reagent is the limited stability of the fluorescent 1,2-disubstituted isoindole.

Effect of impaired renal function on tamoxifen.

There is no information available in the literature on the blood levels of tamoxifen in patients with decreased renal function. As serious side effects of tamoxifen administered at high doses have been reported, a patient with decreased renal function and metastatic breast cancer was studied to determine the blood levels of tamoxifen while under therapy. Since no abnormally elevated levels of tamoxifen were found in this patient during the month of therapy, results of this study indicate that tamoxifen can be administered to patients with some degree of renal impairment without the risk of giving rise to abnormally elevated blood levels.

Influence of tamoxifen and its N-desmethyl and 4-hydroxy metabolites on rat liver microsomal enzymes.

Tamoxifen (Nolvadex; TAM) and its major metabolites, N-desmethyl- (DMT) and 4-hydroxy-tamoxifen (HT), were shown to be potent inhibitors of hepatic cytochrome P-450-dependent mixed function oxidations. From in vitro experiments, all three were found to be potent inhibitors of oxidation of Type-I substrates (ethylmorphine and aminopyrine) and less potent, non-competitive inhibitors of Type-II substrates (aniline and dimethylnitrosamine). TAM, DMT and HT were of essentially equal potency and had a much more pronounced effect on Type-I substrates than on Type-II compounds studied.

Factors affecting the stability of fluorescent isoindoles derived from reaction of o-phthalaldehyde and hydroxyalkylthiols with primary amines.

The stability of a series of fluorescent isoindole derivatives formed in situ under analytical conditions following the reaction of o-phthalaldehyde (OPA) and 2-mercaptoethanol (2-ME) with a series of primary amines are reported. Increasing the bulk and degree of substitution at C-10 of the resulting isoindole resulted in substantial increases in product stability. The effects of excess OPA and 2-ME on isoindole stability were examined and OPA was observed to catalyze isoindole degradation while 2-ME had no effect.

Analysis of riboxamide in plasma by high-performance liquid chromatography using automated column switching.

A sensitive and highly specific assay for riboxamide (TCAR) in human and canine plasma is described. The specificity of the procedure is derived from the method of sample preparation and a high-performance liquid chromatographic separation which utilizes the different selectivities of two columns. Partial separation of TCAR from plasma is achieved on a solvent-generated anion exchanger with silica gel as the solid support.

Tumor concentration of platinum in patients with head and neck cancer.

Simultaneous concentrations of total plasma platinum, filterable platinum, intact cisplatin, and total tumor platinum were measured for 24 hours after an intravenous bolus of cisplatin in five patients. Tumor concentrations of drug were greater than could be explained on the basis of circulating plasma platinum at the time of biopsy. Intratumor platinum concentrations derived from this study provide guidance for selection of the appropriate drug concentrations for in vitro chemosensitivity testing of head and neck cancer in humans.

Evaluation of reductive amperometric detection in the liquid chromatographic determination of antineoplastic platinum complexes.

The usefulness of reductive electrochemical detection at mercury drop electrodes has been determined for platinum complexes separated by solvent-generated anion-exchange high-performance liquid chromatography. Both current-sampled dropping mercury and hanging mercury drop electrodes (DME and HMDE) provide significant advantages over UV absorbance and off-line non-flame atomic absorption detection. The effects of chromatographic and polarographic parameters on analytical system performance have been investigated.

Monitoring the reactions of cisplatin with nucleotides and methionine by reversed-phase high-performance liquid chromatography using cationic and anionic pairing ions.

Methodology, based on reversed-phase high-performance liquid chromatography, is described for monitoring the reactions of cisplatin with DNA, nucleotides, and methionine. Cisplatin was determined in DNA ultrafiltrates on solvent-generated anion exchangers which were prepared by coating the surface of a reversed-phase column with hexadecyltrimethylammonium bromide. These systems were also applicable to studies on the reactions of cisplatin with nucleotides.