Biomedical subjects

D T Rossi

Publications and source records attributed to D T Rossi.

29 records · Page 2Linked to original sources

PubMed1997

Management of nonmatrix interfering peaks in a chiral high-performance liquid chromatographic assay produced by solid-phase extraction of rat plasma.

PD 146923, under evaluation as an alkylating radiosensitizing drug, contains one chiral center and one chemically reactive aziridine ring. A method was developed to evaluate possible in vivo enantiomeric inversion of PD 146923 in rat plasma. Normal-phase chiral HPLC was necessary to separate the enantiomers, but a typical aqueous-based solid-phase extraction (SPE) was needed to isolate the analytes from plasma. SPE at higher analyte concentrations removed all interfering peaks and gave acceptable recoveries. However, peaks (A-G) from seven new components interfering with analyte detection at lower concentrations were produced by SPE. The interfering peaks overlapped each other, so some were not observed until other, more intense interfering peaks had been managed. The low separation efficiency of the chiral column precluded management of interfering peaks by modifying chromatographic parameters. Chemical reactivity of the analytes forced the use of mild conditions for management of interfering peaks. Peaks A-F were: (A) water from the SPE cartridge; (B) SPE sorbent endcapping; (C, E and F) nonvolatile salts of the SPE elution acid reacting with bases from the injection solvent or with unidentified bases from the SPE cartridge; (D and G) analyte degradation products. This study identifies the nonmatrix peaks coeluting with the analytes, and describes how an aqueous-based SPE method was developed for isolating these very polar, highly reactive analytes in plasma for separation in a normal-phase chiral HPLC assay. Additionally, B, C, E or F probably are present in many other solid-phase extractions, but are not observed because of polarity or solubility properties.

Acetic Acid↗

PubMed1997

Analytical considerations for trace determinations of drugs in breast milk.

Recent scientific and regulatory interest in lacteal excretion of drugs has prompted this review of bioanalytical sample preparation techniques for milk. The composition and properties of milk are reviewed, with emphasis on how the sample preparation is affected. The most important principals of mammary gland pharmacology, including protein binding, ion trapping and liquid solubility, are described. Because adequate milk volume is difficult to obtain from some smaller rodent species, special arrangements for sample collection, control preparation and assay standardization often need to be made. Several commonly-used sample preparation approaches for drugs in milk, including direct injection, dialysis and ultrafiltration, protein precipitation, liquid-liquid extraction, solid-phase extraction and immunoaffinity extraction. Have been reported with varying degrees of success. The advantages and disadvantages of each of these approaches is discussed.

Animals↗

PubMed1997

An HPLC assay utilizing solid-phase extraction for CI-1010, an alkylating radiosensitizer, in rat plasma.

CI-1010, a 2-nitroimidazole, is a chiral prodrug for the active moiety PD 146923 and is under development as an alkylating radiosensitizer to be used as an adjuvant to radiotherapy. Because CI-1010 has an estimated half-life < or = 2 min under physiological conditions its metabolites/degradation products PD 146415, an inactive moiety, and PD 146923 were assayed to support rat toxicology studies. The method involves the processing of plasma samples through phenyl solid-phase extraction cartridges followed by chromatography on CN columns with UV detection at 325 nm. The assay appears linear over the range 0.050-100 micrograms ml-1 for both PD 146415 and PD 146923. Interrun accuracy and precision estimates for PD 146415 and PD 146923 were within +/- 6.50 and < or = 3.27%, respectively, and +/- 12.8 and < or = 4.06%, respectively, for quality controls containing nominal concentrations of 0.400, 4.00 and 40.0 micrograms ml-1. The absolute recovery of CI-1010, PD 146415 and internal standard, PD 126675, were approximately 40, 96 and 95%, respectively. The recovery of PD 146923 appeared concentration dependent and ranged from 68 to 92%. PD 146415 and PD 146923 were both stable in rat plasma at 4 degrees C and -77 degrees C for at least 7 h and 154 days, respectively. CI-1010 was not stable in rat plasma at 4 degrees C. CI-1010, PD 146415 and PD 126675 were stable for at least 63 days in 10 mM phosphate buffer at pH 3.0 and 4 degrees C. Under identical conditions PD 146923 was stable for only 8 days. The applicability of this method to determine concentrations of PD 146415 and PD 146923 in rat plasma is reported in this paper.

Alkylating Agents↗

PubMed1996

Design and evaluation of an automated solid-phase extraction method development system for use with biological fluids.

An automated solid-phase extraction method development system, utilizing a Zymate XP robot and a custom-designed solid-phase extraction manifold, has been developed and validated. This system spikes blank liquid matrix, such as plasma, with solutions containing drug, internal standard, and up to three metabolites. Samples are then buffered or diluted with an appropriate reagent. After these samples and corresponding blanks have been prepared, solid-phase cartridges containing selected sorbents are automatically conditioned. Samples are robotically vortexed and transferred to the conditioned cartridges, and analytes are extracted. Validation of this robotic system demonstrated acceptable precision and accuracy for three types of liquid transfer, including metering pump ( < 6% RSD and RE for > or = 2.0 mL dispensation), syringe-based laboratory station ( < or = 2.9% RSD and 0.5% RE for volumes between 0.25 and 1.00 mL), and syringe hands ( < 3.5% RSD and RE for volumes between 0.10 and 1.00 mL). For two example compounds, the system provided data which effectively distinguished good solid-phase sorbents from marginal ones through precision, recovery, and chromatographic selectivity. Solid-phase extraction of these compounds from human plasma gave precision (2-10% RSD) and extraction efficiency (96 +/- 6%) comparable to results obtained from manual extractions (92 +/- 11%).

Body Fluids↗

PubMed1996

Liquid chromatographic assay for a butenolide endothelin antagonist (PD 156707) in plasma.

A sensitive and selective liquid chromatographic assay for determining the non-peptide endothelin A receptor antagonist PD 156707 (I) in rat plasma has been developed and validated. The analyte was isolated from matrix by solid-phase extraction. Liquid chromatographic separation was achieved isocratically on a 3.2 mm I.D., ODS column with a mobile phase of acetonitrile-ammonium phosphate (50 mM, pH 3.5) (44:56, v/v). Column effluent was monitored fluorometrically. Peak-height ratios (analyte/IS) were proportional to I concentrations in rat plasma from 25 to 1000 ng/ml. Assay precision and accuracy for I, based on quality controls, was 9.5% relative standard deviation, with relative error of +/- 6.5%. The quantitation limit was 25 ng/ml for a 200-microliters sample aliquot.

Animals↗

PubMed1995

Trace quantitation of the novel cholinesterase inhibitor in human plasma by capillary gas chromatography/ion trap mass spectrometry.

This paper demonstrates the utility of an ion trap mass spectrometer as a detector for trace quantitative determinations of pharmaceuticals in human plasma by capillary gas chromatography/mass spectrometry. A novel acetylcholinesterase inhibitor (CI-1002) was selected as an illustrative example for the technique. When coupled with a selective solid-phase extraction, this approach was capable of quantifying as little as 34 pg (0.50 ng ml-1, RSD = 12.7%) of compound on the column, and the inter-run precision was typically 3-4% RSD over a 0.5-25 ng ml-1 linear range. The advantages and requirements of the technique, in addition to the prospects for improvements in the detection limit, are discussed.

Calibration↗

PubMed1995

Considerations in the development of a sensitive HPLC assay for human epidermal growth factors in human plasma.

A sensitive assay was developed for human epidermal growth factors (hEGF) 1-48 (dosed), hEGF 1-53 (endogenous), without interference from potential metabolites hEGFs 1-47 or 1-46. Spiked human plasma samples were injected directly, utilizing on-line immunoaffinity HPLC (anti-hEGF) clean-up. No change in capacity was noted after 81 cycles. After release from the immunoaffinity column, the fragments were further resolved by strong cation-exchange (SCX) via a column switching valve. Method development also required interfacing immunoaffinity, ion-exchange, and detection components. Immunoassays on collected fractions yielded a detection limit of 1 microgram ml-1, although a detection limit of 75 pg ml-1 appears feasible.

Amino Acid Sequence↗

PubMed1993

Concentration-dependent protein binding of a novel oral thromboxane synthase inhibitor--FCE 22,178.

The protein binding of FCE 22,178 in humans was determined ex vivo by equilibrium dialysis using plasma samples obtained from a dose-ranging study in normal male volunteers. These data suggested that FCE 22,178 may exhibit concentration-dependent protein binding over an in vivo concentration range of .8 to 64 micrograms/mL. Increase in free fraction at higher plasma drug concentrations corresponded directly to the dose-dependent increase in renal drug clearance. Nonlinear parameter estimation showed that FCE 22,178 binds tightly to plasma proteins with an apparent equilibrium association constant of 1.44 x 10(5) mol/L. Predicted change in the free fraction is consistent with the observed changes in renal clearance.

Blood Proteins↗

PubMed1993

A phase I dose-ranging safety and pharmacokinetics study of a novel oral thromboxane synthase inhibitor, FCE 22, 178.

A 100- to 3200-mg dose range of FCE 22,178 was studied in this phase I single-dose escalation safety/kinetics study. After oral administration, a rapid drug absorptive phase and a biexponential disposition profile were observed. Mean estimates of the terminal elimination half-life of FCE 22,178, over the doses studied, ranged from 7.6 to 14.4 hours. A disproportionate increase in both maximum peak plasma concentration (Cmax) and area under the curve (AUC0-infinity) was noticed for doses higher than 400 mg. Mean estimates of systemic clearance (CLs/F) over the 100- to 400-mg doses were 0.053 to 0.064 L/hour/kg, and were significantly higher for the three higher dose levels. This nonlinearity appears to be related to the changes in oral bioavailability. Estimates of distribution volume (Vd, lambda z/F) for FCE 22,178 increased from 0.75 L/kg at the 100-mg dose to 3.00 L/kg at the 3200-mg dose, and renal clearance (CLr) also increased with dose. Both observations may be related to an increase in free fraction of FCE 22,178 at higher doses. Urinary excretion of unchanged drug averaged < 10% for all dose levels. The urinary excretion of the glucuronide metabolite (M1) averaged 41 to 70% for doses up to 400 mg, but diminished to 13% at the 3200-mg dose. The disposition of M1 appeared to be formation-rate limited. In addition, the ratio of the formation to the disposition clearance for M1 was relatively stable and apparently dose independent. No drug-related adverse experiences were observed over the studied dose range after single doses at FCE 22,178.

Administration, Oral↗

PubMed1992

A sensitive and specific procedure for quantitation of ADR-529 in biological fluids by high-performance liquid chromatography (HPLC) with column switching and amperometric detection.

An HPLC method using electrochemical detection (ED) has been validated for the determination of ADR-529 in plasma and urine using ICRF-192 as an internal standard (IS). Prior to storage and quantitation, both plasma and urine samples require acid stabilization. Acidified plasma samples were prepared for HPLC using a two column solid-phase extraction (SPE). An aliquot of buffered plasma (i.e., pH 6-7) was first deproteinated and desalted on a C-18 SPE column. The analytes were then eluted onto a C-8 SPE column where retention and selective cleanup were achieved in the cation-exchange mode via silanol interactions. Acidified urine samples were diluted in acetonitrile prior to injection. The HPLC system for plasma and urine samples employed two narrow-bore silica columns used in the weak cation-exchange mode and separated by a switching valve. To prohibit late-eluting peaks from passivating the glassy carbon working electrode, a heart-cut containing ADR-529 and the IS was vented from the first silica column to the second using an automated switching valve. Amperometric detection at an oxidation potential of +1050 mV vs a Ag/AgNO3 reference electrode was used. Linearity was validated between 5 and 500 ng/ml in plasma and between 2 and 100 micrograms/ml in urine. Imprecision and percentage bias were typically less than 10% for both plasma and urine controls throughout their respective dynamic ranges. The absolute recoveries for ADR-529 and the IS from plasma were greater than 95%. This method is being successfully applied to the pharmacokinetic/dynamic evaluation of ADR-529 in animals and humans.

Chromatography, High Pressure Liquid↗

PubMed1991

Quantitation of a novel antiemetic (ADR-851) in plasma and urine by reversed-phase high-performance liquid chromatography with fluorescence detection.

A sensitive and specific bioanalytical method for quantitation of a novel antiemetic (ADR-851) in plasma and urine has been developed and validated. The drug and internal standard (metoclopramide) are extracted from the plasma matrix by solid-phase extraction on cyanopropyl bonded-phase columns. After extraction, samples are separated by isocratic reversed-phase high-performance liquid chromatography. The parent drug, internal standard and a yet unidentified metabolite are detected by fluorescence. The method requires 1.0 ml of plasma or 0.1 ml of urine and has a lower limit of quantitation of 2 ng/ml with 10.9% relative standard deviation (R.S.D.). Method linearity has been established over a 2-800 ng/ml range when 1.0 ml of plasma is used. The intra- and inter-day imprecisions for the method are typically better than 6% and 11% R.S.D., respectively, in both plasma and urine over the entire dynamic range. The pooled estimate of bias is less than 5% and attests to the excellent accuracy.

Antiemetics↗