Biomedical subjects

C E Lunte

Publications and source records attributed to C E Lunte.

At least 55 records · Page 3Linked to original sources

PubMed1994

On-line coupling of in vivo microdialysis sampling with capillary electrophoresis.

Microdialysis sampling has become an important means of continuously monitoring reactions in vivo. This sampling technique places a constraint on the analysis method because of the very small sample volume provided. On the other hand, microdialysis provides the advantage of clean samples that do not require cleanup prior to analysis. An on-line coupling of microdialysis sampling to capillary electrophoretic (CE) analysis is described that uses the advantages of microcolumn separations to overcome the small volume limitation. An interface was designed which converts the continuous microdialysis sample stream into discrete 60-nL sample plugs and then injects a portion of this plug into the CE system. The on-line interface provided precision of 2.6% with minimal band broadening or peak height loss relative to off-line sampling. Using a high-speed micellar electrokinetic chromatography (MEKC) separation, resolution of the investigational antineoplastic SR 4233 from its main metabolite SR 4317 was achieved in less than 60 s. This allowed the on-line system to achieve a 90-s temporal resolution for determining the pharmacokinetics of SR 4233 in vivo.

Animals↗

PubMed1994

Microdialysis sampling in tumor and muscle: study of the disposition of 3-amino-1,2,4-benzotriazine-1,4-di-N-oxide (SR 4233).

Microdialysis sampling was used to study the pharmacokinetics and disposition of SR 4233 (3-amino-1,2,4-benzotriazine-1,4-di-N-oxide), a representative of a new class of bioreductive antineoplastic agents. The pharmacokinetics of unbound SR 4233 in plasma was determined using a flexible microdialysis probe implanted into the jugular vein of conscious, freely-moving rats. No difference in the plasma pharmacokinetics was observed between healthy and tumor bearing rats. In both cases a biexponential decay following an i.v. dose was observed with t1/2 (alpha) of 13.4 +/- 3.2 minutes and t1/2 (beta) of 37.0 +/- 14.4 minutes. Binding of SR 4233 to plasma proteins was determined to be concentration independent at 21.4 +/- 2.9%. A linear microdialysis probe was used to sample solid tumor and muscle tissue in vivo to study the disposition of SR 4233 into these tissues. Similar elimination kinetics were observed in both tissues although the peak concentration of SR 4233 in muscle was 20 times that in the tumor. The extent of reductive metabolism was much greater in the tumor relative to muscle. This was observed after both a systemic dose of SR 4233 and direct dosing to the tissue through the microdialysis probe. This study demonstrates the potential of microdialysis sampling in vivo to the study of the disposition of drugs.

Animals↗

PubMed1994

Electrochemistry of catechol-containing flavonoids.

The electrochemical properties of four structurally related flavonoids, quercetin, quercetin-3-O-rhamnose (quercitrin), quercetin-3-O-rutinose (rutin) and luteolin were investigated. These flavonoids were shown to undergo homogenous chemical reactions following oxidation at a glassy carbon electrode. These reactions were studied using cyclic voltammetry and rotating ring-disk voltammetry. Both first-order and zero-order processes were observed. The rate of the zero-order process was strongly dependent on the substituent at the C-3 position of the flavonoid. The rate of the first-order process was independent of substitution. Two products were observed using liquid chromatography. These products did not correspond to previously reported products of enzymatic oxidation. The products were not stable under conditions for isolation.

Electrochemistry↗

PubMed1994

Relationship of flavonoid oxidation potential and effect on rat hepatic microsomal metabolism of benzene and phenol.

The effect of several flavonoids on the benzene hydroxylase and phenol hydroxylase activity of rat hepatic microsomes was determined. The electrochemical characteristics of the flavonoids were also determined. The effect of flavonoids on microsomal phenol hydroxylase activity was found to correlate well with the oxidation potential for flavonoid aglycones. Easily oxidized flavonoids inhibited phenol hydroxylase activity with the extent of inhibition correlated to the ease of oxidation. This inhibition exhibited dose-dependent behaviour, with concentrations below 1 microM having no effect. On the other hand, flavonoids with high oxidation potentials increased phenol hydroxylase activity in a dose-independent manner. Hydroxyl substitution at C-7 was required for inhibition of phenol hydroxylase activity independently of the oxidation potential. Glycosylation at either C-7 or C-3 was found to moderate the inhibition of phenol hydroxylase activity. A linear relation was found between the degree of inhibition and the number of sugar residues for glycosylated flavonoids. There was no correlation between electrochemical properties and effect on microsomal benzene hydroxylase activity.

Animals↗

PubMed1994

Dermal microdialysis sampling in vivo.

Microdialysis sampling of the dermis in vivo was accomplished using a linear microdialysis probe. In contrast to previous studies using a commercial cannula-style microdialysis probe, the linear probe had no effect on the flux of drug through the skin in vitro. The extent of tissue damage in vivo due to probe implantation was evaluated by histological examination and microdialysis delivery studies. Tissue damage due to implantation of the linear probe was minimal with no bleeding or edema observed. Infiltration of lymphocytes into the tissue was observed beginning 6 hours after probe implantation with scar tissue beginning to form after approximately 32 hours. The infiltration of lymphocytes had no effect on the behavior of implanted microdialysis probes. Delivery of 5-fluorouracil was between 20 and 25% for six different probes implanted in six different animals demonstrating good probe-to-probe and implantation-to-implantation reproducibility. Constant delivery was maintained for at least 24 hours in all cases indicating that experiments of at least 24 hour duration are feasible. The dermal concentration of topically applied 5-FU cream, Efudex, was continuously monitored by an implanted microdialysis probe demonstrating the feasibility of this technique as for monitoring skin drug levels in vivo. The dermal concentration of 5-FU following topical application was approximately 40-fold higher for in vitro excised skin than for in vivo intact skin.

Animals↗

PubMed1993

Examination of microdialysis sampling in a well-characterized hydrodynamic system.

The use of microdialysis sampling was examined in a well-characterized hydrodynamic system. A cross-flow microdialysis probe was designed in which the flow of both the dialysis perfusion solution and the sample solution could be carefully controlled. Dialysis membranes of cellulose (Cuprophan), cellulose acetate, and polyacrylonitrile (PAN) were examined in this system using hydroquinone as the test analyte. The permeability of the membranes to hydroquinone ranged from 1.72 x 10(-6) cm2/s for PAN to 2.97 x 10(-7) cm2/s for cellulose acetate. Determination of the dialysis fibers' recovery as a function of the sample flow velocity resulted in a rapid increase in recovery with increase in flow velocity. The recovery plateaued at high sample velocity. These results show that at low sample velocity diffusion through the sample solution is the rate-limiting step in recovery while at higher velocity transport through the membrane becomes rate limiting. Recovery for all three membrane types plateaued above sample velocities of 0.211 cm/s. This is well below the velocity of most biological fluids in which microdialysis sampling has been applied. This result supports previous reports that an in vitro calibration of microdialysis probes is appropriate for use in hydrodynamic environments in vivo such as the blood and bile.

Chromatography, Liquid↗

PubMed1993

Enzymatic formation and electrochemical characterization of multiply substituted glutathione conjugates of hydroquinone.

1,4-Benzoquinone can undergo redox cycling in the presence of glutathione to produce multiply substituted products. It has previously been shown that the nephrotoxicity of the hydroquinone-glutathione conjugates increases with increasing substitution. However, based on chromatographically-assisted hydrodynamic voltammetry (CA-HDV), the oxidation potential was shown to apparently increase which, should lead to decreased toxicity. From the chemical formation of multiply substituted hydroquinone-glutathione conjugates from benzoquinone and glutathione, it is clear that the thermodynamic oxidation potential must decrease as substitution increases. This was confirmed by cyclic voltammetric (CV) characterization of the isolated conjugates. The discrepancy between the CV and CA-HDV data apparently results from kinetic factors arising from differences in the treatment of the electrode surface between the two experiments. The multiply substituted hydroquinone-glutathione conjugates were also produced in horseradish peroxidase incubations containing hydroquinone and glutathione. These products were identified chromatographically, spectrophotometrically, and electrochemically. The increasing ease of oxidation and the possible enzymatic formation of multiply substituted hydroquinone-glutathione conjugates indicates that this pathway may occur in vivo and contribute to the toxicity of quinones.

Chromatography, Liquid↗

PubMed1993

Distribution of tacrine across the blood-brain barrier in awake, freely moving rats using in vivo microdialysis sampling.

Microdialysis was used to sample simultaneously the distribution of THA (9-amino-1,2,3,4-tetrahydroacridine; Tacrine), a potential anti-Alzheimer agent, both in blood and across the blood-brain barrier of anesthetized and awake, freely moving rats. Microdialysis probes were implanted in the jugular vein and dorsal hippocampus and dialysis samples were simultaneously collected from both sites. Dialysis samples were analyzed using a microbore column chromatographic assay with a detection limit of 0.3 ng/ml. Pharmacokinetic parameters were calculated after a 1 mg/kg intravenous dose of THA. Plasma pharmacokinetics followed a biexponential mode, with t1/2(dis.) = 8.4 +/- 2.7 min and t1/2(elim.) = 76.7 +/- 24.2 min for awake, freely moving rats. THA rapidly penetrated the blood-brain barrier, with maximum concentrations attained within 60 min post-dose. In the brain of awake, freely moving rats t1/2(abs.) was 26.0 +/- 5.2 min and t1/2(elim.) was 99.1 +/- 17.7 min. THA levels in hippocampus extracellular fluid were 10 times lower than those in plasma. For anesthetized rats, the t1/2(elim.) in blood was 154.8 +/- 46.8 min, while in the hippocampus t1/2(elim.) was 159.5 +/- 31.7 min. The binding of THA in rat plasma was 56.2 +/- 5.0%, while the fraction bound to rat whole blood was 73.3 +/- 4.1% as determined by microdialysis and ultrafiltration.

Animals↗

PubMed1993

In vivo microdialysis sampling in the bile, blood, and liver of rats to study the disposition of phenol.

Methods for continuous in vivo sampling in the bile, blood, and liver extracellular fluid are described. These methods are based on microdialysis sampling in anesthetized rats. A new flow-through microdialysis probe is described for sampling bile while maintaining normal bile flow. All three sites are simultaneously and continuously sampled to provide concentration-time profiles at multiple sites in a single experimental animal. This technique is demonstrated by studying the hepatic metabolism and biliary excretion of phenol in rats. Following an i.v. infusion of phenol, the major hepatic metabolite was found to be phenyl-glucuronide. Hydroquinone and 2-glutathionyl-hydroquinone were also detected but at lower concentrations. A similar pattern of metabolites was found in the bile and blood. For all of the metabolites, bile concentrations are higher than liver concentrations, indicating that the metabolites are actively excreted into the bile.

Animals↗

PubMed1992

Monitoring excitatory amino acid release in vivo by microdialysis with capillary electrophoresis-electrochemistry.

Capillary electrophoresis (CE) with electrochemical detection (ED) was used to determine extracellular levels of aspartate, glutamate and alanine in samples from the frontoparietal cortex of the rat which were obtained by microdialysis. The method was used to monitor the effect on the overflow of the excitatory amino acids aspartate and glutamate of an influx of high concentrations of potassium ion. Samples were derivatized with naphthalenedialdehyde-cyanide prior to analysis. Detection limits for aspartate and glutamate were 80 and 100 nM, respectively. CE-ED is extremely useful for the analysis of microdialysis samples because of the very small sample volumes required by this analytical technique. The use of ED provides the requisite sensitivity and allows verification of peak purity by voltammetry.

Amino Acids↗

PubMed1992

Intravenous microdialysis sampling in awake, freely-moving rats.

Intravenous microdialysis sampling in the awake, freely-moving rat for the determination of free drug concentrations in blood is described. Intravenous microdialysis was performed with a nonmetallic, flexible dialysis probe. The pharmacokinetics of theophylline were determined using both microdialysis sampling and collection of whole blood following an iv dose. There was no difference in the half-life of elimination of theophylline determined by microdialysis, 4.4 +/- 0.4 h, and whole blood sampling, 4.5 +/- 0.7 h. The kinetics of elimination were affected by removing blood samples and by using anesthesia. The half-life of elimination was 4.4 +/- 0.4 h when using simultaneous microdialysis and whole-blood sampling and only 3.0 +/- 0.4 h using microdialysis alone. The half-life of elimination was 17.0 +/- 7.1 h in chloral hydrate anesthesized rats. Microdialysis samples were continuously collected for over 7 h without fluid loss using a single experimental animal. Microdialysis sampling directly assesses the free drug concentration in blood. The extent of theophylline binding to blood proteins was determined in vitro in rat plasma and rat whole blood using both ultrafiltration and microdialysis. Theophylline was (47.3 +/- 1.3)% bound in rat plasma and (52.2 +/- 1.6)% bound in rat whole blood. Microdialysis sampling is a powerful tool for pharmacokinetic studies, providing accurate and precise pharmacokinetic data.

Anesthesia↗

PubMed1992

Investigation of the degradation mechanism of 5-aminosalicylic acid in aqueous solution.

The solution degradation of the antiinflammatory agent 5-aminosalicylic acid (5-ASA) was investigated in order to elucidate a mechanism for degradation. Two degradation pathways were considered: decarboxylation by analogy to 4-aminosalicylic acid (4-ASA) decomposition and oxidation from consideration of 5-ASA's aromatic ring substitution pattern (i.e., relation to p-aminophenol). The oxidation of 5-ASA was investigated using cyclic voltammetry and flow electrolysis. These studies showed that 5-ASA is more easily oxidized than is 4-ASA and that 5-ASA undergoes a two-electron, two-proton oxidation consistent with formation of 5-ASA-quinoneimine (5-ASA-QI). This oxidation is followed by subsequent complex chemistry. The decomposition of 5-ASA in solution was examined under a variety of conditions. 5-ASA decomposes most rapidly under conditions promoting oxidation and is most stable under conditions tending to inhibit oxidation. Decarboxylation was not found to be a significant degradation pathway.

Aminosalicylic Acid↗

PubMed1992

Microdialysis sampling for the investigation of dermal drug transport.

Microdialysis perfusion in vivo has the potential to be a powerful sampling technique in dermal and transdermal drug delivery studies. Characterization of a commercially available microdialysis probe in vitro considering relevant physiological parameters is a vital first step in the evaluation of microdialysis as a dermal sampling technique. In previous microdialysis studies, analyte concentration and neutrality have been implicated in altering microdialysis recovery. The recovery of a model compound 5-fluorouracil (5-FU) was investigated at several pH values and donor concentrations. The relative recovery of 5-FU by the microdialysis probe was affected by pH but not by donor concentration. To confirm further that the changing concentration and pH profile presented by the flux of 5-FU was not significantly altering microdialysis recovery, an experiment comparing direct and microdialysis sampling of a Franz diffusion cell receptor compartment was performed. Although the 5-FU concentration (0-686 ng/ml) and pH (7.40-7.24) changed substantially, the recovery of 5-FU was not adversely affected. To demonstrate the feasibility of dermal microdialysis, the flux of a commercial preparation of 5-fluorouracil was monitored utilizing a microdialysis probe implanted in excised rat skin in vitro. The results from the dermally implanted probe demonstrate the potential of the technique while establishing the limitations of the current microdialysis system.

Animals↗

PubMed1991

In vivo microdialysis sampling for pharmacokinetic investigations.

In vivo microdialysis sampling coupled to liquid chromatography was used to study acetaminophen disposition in anesthetized rats. The pharmacokinetics of acetaminophen and its sulfate and glucuronide metabolites were determined using both microdialysis sampling and collection of whole blood. For microdialysis, samples were continuously collected for over 5 hr without fluid loss using a single experimental animal. Microdialysis sampling directly assesses the free drug concentration in blood. The pharmacokinetic results obtained with microdialysis sampling were the same as those obtained from blood collection. The administration of heparin, necessary when collecting blood samples, was found to double the elimination half-life of acetaminophen. Microdialysis sampling is a powerful tool for pharmacokinetic studies, providing accurate and precise pharmacokinetic data.

Acetaminophen↗

PubMed1990

In vivo microdialysis sampling coupled to liquid chromatography for the study of acetaminophen metabolism.

In vivo microdialysis sampling coupled to liquid chromatography is a powerful tool for the study of drug metabolism. This technique is illustrated by investigating the pharmacokinetics of acetaminophen in the blood and liver of an anesthetized rat. The pharmacokinetics of the sulfate and glucuronide metabolites as well as the parent acetaminophen can be determined with high precision using microdialysis sampling. Microdialysis samples can be collected at a high rate from several sites without fluid loss with a single animal. Because the animal serves as its own control better data can be obtained. Liquid chromatography provides determination of multiple analytes per sample for metabolic profiling. This technique will provide more accurate and precise pharmacokinetic data while requiring fewer animals.

Acetaminophen↗

PubMed1990

Detection of N-acetylcysteine, cysteine and their disulfides in urine by liquid chromatography with a dual-electrode amperometric detector.

A method for the determination of N-acetylcysteine, cysteine and their disulfides in urine is described. The thiols and disulfides are separated by reversed-phase ion-pair chromatography with octyl sodium sulfate as the ion-pairing reagent and detected with a dual-electrode amperometric detector using Au/Hg amalgam electrodes. Both the thiols and disulfides are detected with this system. In addition, dimers and mixed disulfides can be detected individually.

Acetylcysteine↗

PubMed1990

Electroanalytical approaches to understanding benzene metabolism.

Electrochemical techniques are ideally suited to the study of the metabolism of aromatic xenobiotics because the metabolites are frequently easier to oxidize than the parent compounds. In many cases, the trace metabolites have the lowest oxidation potentials and hence electrochemical methods have the greatest selectivity for these compounds. The sensitivity of dual-electrode liquid chromatography-electrochemistry for the detection and identification of trace metabolites was demonstrated by the detection of the secondary metabolite, hydroquinone, in a microsomal incubation containing benzene and ascorbic acid. The use of an electrochemical detector in a series configuration provides increased selectivity for chemically reversible metabolites such as hydroquinone. Electrochemical methods can also be used to generate metabolites. The products of the electrochemical oxidation of phenol and biphenol are compared with those generated in a peroxidase incubation.

Animals↗

PubMed1990

Electrochemical investigations of immunologically reactive procainamide metabolites.

As a result of the implication of N-oxidized procainamide metabolites in drug-related lupus (DRL), the electrochemical behaviour of these compounds was investigated and a coulometric synthesis of the nitroso derivative developed using a previously described carbon packed bed bulk electrolysis flow cell. The electrochemical characterization of the parent p-substituted aromatic amine and the N-oxidized derivatives was achieved through systematic comparison with previously well described aromatic amine and nitro systems using cyclic voltammetry and liquid chromatography with electrochemical detection (LC-EC). Chromatographically assisted hydrodynamic voltammetry indicated current limiting plateau potentials of 0.45 and -0.2 V versus Ag/AgCl, respectively, for synthetically prepared procainamide hydroxylamine and electrolytically prepared nitrosoprocainamide. Reaction characterization and binding behaviour is described for each of the procainamide metabolites following in vitro incubations with cysteine, glutathione, ascorbic acid and mouse haemoglobin.

Chromatography, High Pressure Liquid↗