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At least 577 records · Page 32Linked to original sources
PubMed1995

Effects of 9-aminocamptothecin on newly synthesized DNA in patient bone marrow samples.

9-Aminocamptothecin (9-AC) inhibited cell growth and DNA synthesis in HCT 116 human colon cancer cells in a concentration- and time-dependent manner. Interference with nascent DNA chain elongation was monitored using pH step alkaline elution. After a 3-day 9-AC exposure, 38% (10 nM) and 53% (50 nM) of the total [3H]DNA eluted with pH steps 11.3-11.7, compared to 9% in control cells. Effects on nascent DNA integrity were also evaluated by fixed elution with pH 12.1 buffer. After a 3-day exposure to 9-AC, 27% (10 nM) and 82.5% (50 nM) of the total [3H]DNA eluted relative to control. Paired bone marrow samples were then obtained in 10 patients before treatment and between 42 and 72 h of a continuous i. v. infusion of 9-AC (35-74 microgram/m2/h for 72 h). The mononuclear cells were incubated with [3H]dThd for 2 or 4 h, and then analyzed using either pH step or fixed pH alkaline elution, respectively. In seven patients receiving >/=47 microgram/m2/h 9-AC, 4% +/- 1.5% (mean +/- SE) of the total [3H]DNA eluted with pH steps </= 11.7 in the pretreatment samples compared to 13% +/- 3.6% during 9-AC (P = 0. 037). An altered fixed pH elution profile of nascent DNA was noted in two patients treated with 59 and 74 microgram/m2/h 9-AC compared to baseline. DNA synthesis was inhibited by 89% +/- 5% during infusion of >/=59 microgram/m2/h 9-AC (n = 7). Since hematological toxicity is dose limiting on this 9-AC schedule, these cellular pharmacodynamic studies provide evidence of a DNA-directed cytotoxic effect of 9-AC in a sensitive host target tissue.

Adult↗
PubMed2005

Emergence of antiretroviral resistance in HIV-positive women receiving combination antiretroviral therapy in pregnancy.

BACKGROUND: Antenatal antiretroviral therapy is integral to preventing vertical transmission of HIV-1. The impact of temporary triple antiretroviral therapy in pregnancy on the emergence of antiretroviral resistance has not been studied. OBJECTIVE: To determine the impact of temporary triple antiretroviral therapy in pregnancy on emergence of antiretroviral resistance. METHODS: Pregnant HIV-1 infected women with a pre-treatment CD4 cell count >300 x 10(6)/l initiated triple antiretroviral therapy in the third trimester and discontinued postpartum. Genotypic resistance testing was performed after antiretroviral cessation and on pretreatment samples when postpartum samples showed primary mutations. RESULTS: In a cohort of 50 women who initiated antiretroviral therapy in pregnancy, 39 (78%) had postpartum HIV-1 nucleotide sequences available for analysis: 35 of these (90%) were previously antiretroviral naive. Seven primary mutations, V106A (one), Y181C (two), G190A (one), K101E (one), M184V (one), T215S (one) were detected in five (13%) women. All five were on regimens that included nevirapine and all were antiretroviral therapy naive prior to the index pregnancy. Four had no mutations detected pretreatment (one did not have a pretreatment analysis available; viral load 83 copies/ml). The median duration of antiretroviral exposure was 70 days. CONCLUSION: Emergence of genotypic resistance is significant in this cohort of pregnant women. All mutations detected were in those that took nevirapine-containing regimens. The clinical implications of these mutations are unknown.

Adolescent↗
PubMed1987

Biogenic amine regulation of bovine luteal progesterone production in vivo.

Biogenic amines were administered using osmotic pumps placed subcutaneously in the neck region of regularly cycling, non-lactating dairy cows on Days 9-11 (oestrus = Day 0) of the oestrous cycle. Blood samples were collected using indwelling jugular catheters and the plasma progesterone concentrations were measured. Samples were collected at 4-h intervals for the first 12 h of treatment and thereafter at 12-h intervals for the remainder of the 72-h treatment period. After administration of various doses of noradrenaline, adrenaline and serotonin (0.5-2.0 micrograms/kg/h) significant elevation of plasma progesterone was achieved at a dosage of 2.0 micrograms/kg/h (P less than 0.01). The response to adrenaline was greater than that observed for noradrenaline and serotonin (P less than 0.05). Within-treatment comparison to pretreatment samples showed plasma progesterone concentrations to increase within 4 h after the administration of noradrenaline, adrenaline and serotonin (P less than 0.05) and this enhancement was maintained throughout the treatment period (P less than 0.05). The elevation in plasma progesterone concentrations induced by noradrenaline, adrenaline and serotonin was independent of changes in circulating concentrations of luteinizing hormone. These results support a physiological role for endogenous biogenic amines in the control of bovine luteal progesterone production.

Animals↗
PubMedDate not supplied

Liquid chromatographic multicomponent method for determination of residues of ipronidazole, ronidazole, and dimetridazole and some relevant metabolites in eggs, plasma, and feces and its use in depletion studies in laying hens.

A simple, rapid liquid chromatographic (LC) method that uses UV/VIS detection has been developed for the determination in eggs of residues of the histomonostats dimetridazole (DMZ), ronidazole (RON), ipronidazole (IPR), and side-chain hydroxylated metabolites of DMZ and RON. Sample pretreatment includes an aqueous extraction, purification with an Extrelut cartridge, and acid partitioning with isooctane. An aliquot of the final aqueous extract is injected into a reverse-phase LC system; detection is performed at 313 nm. The limits of determination are in the 5-10 microgram/kg range. A UV/VIS spectrum can be obtained at the 10 microgram/kg level by using diode-array UV/VIS detection. Recoveries are between 80 and 98% with a coefficient of variation of about 5%. Some 20 samples can be analyzed per day. A side-chain hydroxylated metabolite of IPR can also be detected with this method, as demonstrated with samples from animal experiments. After a single oral dose of the drugs to laying hens, residues of the parent compound and/or the hydroxylated metabolites could be detected in eggs 5-8 days after dosing. Plasma distribution and excretion in feces were established both with and without deconjugation. DMZ and IPR were extensively metabolized to hydroxylated nitroimidazole metabolites; RON was excreted mainly as the parent compound.

Animals↗
PubMed1990

Rapid determination of clenbuterol residues in urine by high-performance liquid chromatography with on-line automated sample processing using immunoaffinity chromatography.

A liquid chromatographic column-switching system for automated sample pretreatment and determination of clenbuterol in calf urine, using an immunoaffinity precolumn with Sepharose-immobilized polyclonal antibodies against clenbuterol, is described. A second precolumn packed with C18-bonded silica was used for the reconcentration of desorbed clenbuterol prior to the analytical separation. Urine, after 2-fold dilution with buffer (pH 7.4), was loaded directly onto the immuno precolumn, where clenbuterol was trapped by the immobilized antibodies. This immuno precolumn has been used for more than 200 runs with standard solutions and samples. Bound analyte was desorbed with 0.01 M acetic acid and transferred, via the second precolumn, to the analytical column. The total runtime per sample was 35 min. Using a sample load of 27 ml of dilute urine and UV detection at 244 nm, the detection limit was 0.5 ng/ml. The mean recovery of clenbuterol added to a blank urine sample at the 5 ng/ml level was 82 +/- 2% (n = 5) as determined with standard solutions loaded onto the same system. Urine samples from treated animals were analysed and the clenbuterol concentrations were comparable to those obtained by high-performance liquid chromatography using solid-phase extraction for sample clean-up.

Animals↗
PubMed1992

Enantioselective determination of R- and S-(alpha-bromoisovaleryl)urea in plasma using high-performance liquid chromatography after solid-phase extraction.

A stereoselective method has been developed for the determination of R- and S-(alpha-bromoisovaleryl)urea in plasma and saliva after oral administration. The chiral separation was carried out on Chiralcel OJ or OD columns with hexane--2-propanol as the mobile phase. The poor detection properties of the analyte required the development of an effective sample pretreatment procedure to enable ultraviolet detection at 210 nm. Solid-phase extraction using hydrophobic Amberlite XAD-2 in combination with washing steps at alkaline and acidic pH completely removed interfering components of the biological matrix and allowed the detection of the optical isomers at concentrations down to 10 ng/ml (0.05 microM). The method was validated by determining the recovery, linearity, accuracy and within-day and between-day repeatability at 50, 200 and 2000 ng/ml. Application to the analysis of plasma and saliva samples is demonstrated.

Bromisovalum↗
PubMed2000

Sequence variations of precore/core and precore promoter regions of hepatitis B virus in patients with or without viral reactivation during cytotoxic chemotherapy.

Reactivation of the hepatitis B virus (HBV) is a well-described complication among cancer patients undergoing cytotoxic chemotherapy. Mutations in the preC/C and the preC promoter regions of HBV have been reported in some patients who developed this condition. A G-to-A mutation at nt 1896 in the preC/C region (HBeAg negative/ anti-HBe positive) has been associated with more severe liver disease than that caused by wild type virus. In addition, it has been suggested that patients with these mutations may be more likely to reactivate than those with the wild type virus. Whether or not such mutations were present before the commencement of or developed during the course of cytotoxic chemotherapy is not known. In this study, 28 cancer patients (consisting of 14 consecutive patients who developed HBV reactivation and another 14 who had no reactivation during cytotoxic chemotherapy) are reported. The objectives were firstly, to determine the prechemotherapy HBeAg status and nucleotide sequences of the preC/C and preC promoter regions of HBV in order to determine if these parameters affected the rate of reactivation, and secondly, for those who developed reactivation, to determine whether the mutations were present before chemotherapy or developed during, possibly as a result of, cytotoxic chemotherapy. HBV DNA was amplified by PCR and nucleotide sequencing performed on samples taken prior to chemotherapy and at the time of reactivation. Results revealed that 16 of the 28 patients were HBeAg negative/anti-HBe positive. Of these 16, four (57%) of the seven patients who had nt 1896 mutation, but only one (17%) of the six who had the wild type HBV genome, developed reactivation. Three had no detectable HBV DNA. In the majority of cases, the type of virus, i.e. wild/mutant at preC/C, that was detected during the reactivation was identical to that detected in the pretreatment samples. With respect to the preC promoter region, the two commonest mutations detected were at nt 1762 (A to T) and nt 1764 (G to A). When this region was translated into amino acid sequences, stop codons leading to truncated X protein at carboxyl terminus were found in four patients, three of whom developed HBV reactivation. We conclude that chronic HBV carriers who are HBeAg negative/anti-HBe positive with nt 1896 mutation (G to A) may be more likely to develop HBV reactivation during cytotoxic chemotherapy than those with the wild type virus. Cytotoxic chemotherapy does not appear to select out mutant HBV, or to be consistently mutagenic in patients who develop HBV reactivation. The occurrence of stop codons in the amino acid sequences of the X protein in three patients who developed HBV reactivation, including one who was detected only at the time of reactivation, is of particular interest, as such mutant viruses remain replication competent.

Adult↗
PubMed2026

Development and clinical validation of a CRISPR/Cas9-engineered reporter phage cocktail for rapid detection of Escherichia coli in urine.

Urinary tract infections are one of the most common infectious diseases, with Escherichia coli as the predominant pathogen. Traditional diagnostic methods fail to meet clinical demands for rapid and specific detection. Here, we developed an efficient urine E. coli detection strategy via a reporter phage cocktail. Four reporter phages (T2::Nluc, T4::Nluc, T5::Nluc, T6::Nluc) were constructed by the CRISPR/Cas9 system combined with homologous recombination. One-step growth curves, optimal multiplicity of infection, and lytic efficiency showed that the Nluc gene block insertion exerted heterogeneous effects on phages. Luminescence assays demonstrated that all five reporter phages (including previously preserved T7::Nluc) and the cocktail offered favorable limits of detection (&#x2265;103 CFU/mL), high specificity, and no urine matrix interference. However, single phages exhibited limited coverage among 177 clinical E. coli isolates. But the reporter phage cocktail remedies this limitation. In large-scale clinical validation, the cocktail achieved sensitivity 73.15% (63.76%-81.22%), specificity 100.00% (99.53%-100.00%), positive predictive value (PPV) 100.00% (95.44%-100.00%), and negative predictive value (NPV) 96.42% (95.18%-97.36%) (all 95% confidence interval [CI]), and excellent concordance with the gold-standard method (Kappa = 0.83, 95% CI: 0.77-0.89), greatly outperforming single reporter phages (~40.00% sensitivity). This method requires no sample pretreatment, is simple to operate, and completes detection within 4 h, significantly improving diagnostic efficiency. Accordingly, it provides a novel platform for pathogen detection and supports the clinical translation of reporter phage diagnostics.IMPORTANCEUrinary tract infections impose substantial economic and public health burdens. In this study, we successfully constructed Escherichia coli-specific reporter phages T2::Nluc, T4::Nluc, T5::Nluc, and T6::Nluc. Combined with the previously preserved T7::Nluc, these phages formed a reporter phage cocktail. Co-cultivation of this cocktail with clinical samples enabled rapid and specific detection of E. coli in clinical urine, with a significantly shortened detection time (4 h) and good concordance with the gold-standard detection method (Kappa = 0.83), effectively improving detection efficiency and accuracy. This novel pathogen detection platform, integrating specific recognition and signal amplification, not only provides a new technical approach for the rapid and accurate diagnosis of clinical urinary tract infections but also effectively promotes the coordinated improvement of infectious disease diagnosis and treatment in terms of timeliness-precision-cost.

Escherichia coli↗
PubMed2003

Garlic (Allium sativum L.) modulates cytokine expression in lipopolysaccharide-activated human blood thereby inhibiting NF-kappaB activity.

Garlic is proposed to have immunomodulatory and anti-inflammatory properties. This paper shows that garlic powder extracts (GPE) and single garlic metabolites modulate lipopolysaccharide (LPS)-induced cytokine levels in human whole blood. GPE-altered cytokine levels in human blood sample supernatants reduced nuclear factor (NF)-kappaB activity in human cells exposed to these samples. Pretreatment with GPE (100 mg/L) reduced LPS-induced production of proinflammatory cytokines interleukin (IL)-1beta from 15.7 +/- 5.1 to 6.2 +/- 1.2 micro g/L and tumor necrosis factor (TNF)-alpha from 8.8 +/- 2.4 to 3.9 +/- 0.8 micro g/L, respectively, whereas the expression of the anti-inflammatory cytokine IL-10 was unchanged. The garlic metabolite diallydisulfide (1-100 micro mol/L) also significantly reduced IL-1beta and TNF-alpha. Interestingly, exposure of human embryonic kidney cell line (HEK293) cells to GPE-treated blood sample supernatants (10 or 100 mg/L) reduced NF-kappaB activity compared with cells exposed to untreated blood supernatants as measured by a NF-kappaB-driven luciferase reporter gene assay. Blood samples treated with extract obtained from unfertilized garlic (100 mg/L) reduced NF-kappaB activity by 25%, whereas blood samples treated with sulfur-fertilized garlic extracts (100 mg/L) lowered NF-kappaB activity by 41%. In summary, garlic may indeed promote an anti-inflammatory environment by cytokine modulation in human blood that leads to an overall inhibition of NF-kappaB activity in the surrounding tissue.

Cell Line↗
PubMed1998

Determination of illicit and/or abused drugs and compounds of forensic interest in biosamples by capillary electrophoretic/electrokinetic methods.

The application of capillary electrophoresis (CE) methods in forensic toxicology for the determination of illicit and/or misused drugs in biological samples is reviewed in the present paper. Sample pretreatments and direct injection modes used in CE for analysis of drugs in biological fluids are briefly described. Besides, applications of separation methods based on capillary zone electrophoresis or micellar electrokinetic chromatography with UV absorbance detection to (i) analysis of drugs of abuse, (ii) analysis of other drugs and toxicants of potential forensic interest and (iii) for metabolism studies are reviewed. Also, alternative CE methods are briefly discussed, including capillary isotachophoresis and separation on mixed polymer networks. High sensitivity detection methods used for forensic drug analysis in biological samples are then presented, particularly those based on laser induced fluorescence. A glimpse of the first examples of application of CE-mass spectrometry in forensic toxicology is finally given.

Body Fluids↗
PubMedDate not supplied

Determination of available phenolic compounds in soils by liquid chromatography with solid-phase extraction.

A fast, selective, and sensitive liquid chromatographic (LC) method was developed for determination of derivatives of benzoic and cinnamic acids (gallic, protocatechuic, 2,3,4-trihydroxybenzoic, 4-hydroxybenzoic, vanillic, caffeic, syringic, 4-coumaric, ferulic, sinapic, benzoic, 2-coumaric, cinnamic acids, and 4-hydroxybenzaldehyde and vanillin) in soil samples. The method for sample pretreatment is based on temperature-controlled extraction with water (pH 5.6) for 60 min. Extracts were preconcentrated and purified by solid-phase extraction on OASIS HLB sorbent, with subsequent separation and quantification of individual substances by LC with UV diode-array detection. Limits of detection (3 signal-to-noise LODs) better than 65 ng/g (dry weight) and recoveries from 88 to 99% were found for each compound at absorbance 280 nm. The method was used for determination of bioavailable phenolic compounds in different soil samples.

Chromatography, Liquid↗
PubMedDate not supplied

Main component assay of pharmaceuticals by capillary electrophoresis: considerations regarding precision, accuracy, and linearity data.

Capillary electrophoresis has been successfully employed to determine the level of drugs in a variety of pharmaceutical preparations. A large number of reports have shown agreement between CE results and HPLC data or with label claim. Currently the use of CE for main component assays constitutes 26% of the routine usage of CE within drug companies and is the most frequent application. The choice between adopting CE or HPLC for a particular application is very dependent upon the relative merits of each technique to the individual assay. Often, CE can have advantages in terms of reduced sample pretreatment, consumable costs, and analysis time. The ability to separate a wide range of solutes using a single set of operating conditions is a strong advantage of CE. This paper extensively reviews the literature reports of the use of CE for main peak assay and indicates the validation performance data achieved. The specific requirements relating to optimized accuracy and precision in CE assay are discussed in some detail. The use of an appropriate internal standard to improve performance for precision, accuracy, and linearity, and to reduce the impact of sample matrix effects, is experimentally shown by results from the analysis of levothyroxine samples. The applications are subdivided into those samples analyzed by free solution capillary electrophoresis (FSCE) at low or high pH or those separated by micellar electrokinetic capillary electrophoresis (MECC).

Chromatography, High Pressure Liquid↗
PubMed1998

High-performance liquid chromatographic determination of stabilized 4-hydroxyifosfamide in human plasma and erythrocytes.

A method using reversed-phase high-performance liquid chromatography (RP-HPLC) is described for the measurement of the stabilized activated metabolite of ifosfamide, 4-hydroxyifosfamide (4-OHIF), in human plasma and erythrocytes. Immediately after sample collection and plasma-erythrocyte separation at 4 degrees C, 4-OHIF was stabilized by derivatization with semicarbazide (SCZ). The sample pretreatment involved liquid-liquid extraction with ethyl acetate. RP-HPLC was executed with a C8 column and acetonitrile-0.025 M potassium dihydrogenphosphate buffer (pH 7.40)-triethylamine (13.5:86:0.5, v/v) as mobile phase. The analyte was determined with UV detection at 230 nm. Complete validation, optimisation and stability studies were performed and the method proved to be specific, sensitive and with a stable analyte in the range of clinically relevant concentrations (0.1-10 microg/ml) after conventional dosing. The lower limit of quantitation was 100 ng/ml using 1.00 ml of sample. Accuracy was between 94.1 and 107.0%. Within-day and between-day precisions were less than 6.2% and 7.2%, respectively. 4-OHIF-SCZ was found to be stable in the biological matrix at -20 degrees C for at least 1 month. A pharmacokinetic study conducted in a patient receiving 9 g/m2 over 3 days by means of a continuous infusion, demonstrated the applicability of this method.

Acetonitriles↗
PubMed1999

Isotope dilution-gas chromatography/mass spectrometry and liquid chromatography/electrospray ionization-tandem mass spectrometry for the determination of triiodo-L-thyronine in serum.

Isotope dilution-gas chromatography/mass spectrometry (ID-GC/MS) and isotope dilution-liquid chromatography/tandem mass spectrometry (ID-LC/MS/MS) methods have been developed for an determination of triiodothyronine (T3) in serum and their potential as candidate reference methods investigated. In both methods, (13)C(9)-T3 was used as internal standard. Sample pretreatment consisted of deproteinization, extraction and high performance liquid chromatography (HPLC) purification. Conversion of serum thyroxine (T4) to T3 was controlled by adding (13)C(6)-T4. For GC/MS, T3 and (13)C(9)-T3 were converted to the N,O-di-heptafluorobutyryl (HFB) methyl ester derivatives and monitored at m/z 844 and 853. For LC/MS with electrospray ionization, the transitions m/z 652/661 to 606/614 were monitored. For use of the methods as candidate reference methods, special attention was paid to the calibration and the measurement protocol (duplicate analysis of each sample on three occasions). Evaluation of the ID-GC/MS and ID-LC/MS/MS methods showed the absence of interference by reverse T3 and T4, a limit of detection of 100 pg (GC/MS) and 18 pg (LC/MS), a recovery of 100 +/- 1.5% (95% confidence interval) and good precision (the total coefficient of variation, n = 6, was typically 1.5%). In addition to the recovery study, the accuracy of the methods was proven by method comparison (ID-GC/MS vs. LC/MS/MS) on three sera, showing a maximum deviation of 1.1%. Finally, the ID-GC/MS method was applied for measurement of 10 different human sera with a T3 concentration range from 0.6 to 7.3 ng/mL.

Carbon Isotopes↗
PubMed2001

Rapid bioanalysis of vancomycin in serum and urine by high-performance liquid chromatography tandem mass spectrometry using on-line sample extraction and parallel analytical columns.

A novel high-performance liquid chromatography tandem mass spectrometry (LC/MS/MS) method is described for the determination of vancomycin in serum and urine. After the addition of internal standard (teicoplanin), serum and urine samples were directly injected onto an HPLC system consisting of an extraction column and dual analytical columns. The columns are plumbed through two switching valves. A six-port valve directs extraction column effluent either to waste or to an analytical column. A ten-port valve simultaneously permits equilibration of one analytical column while the other is used for sample analysis. Thus, off-line analytical column equilibration time does not require mass spectrometer time, freeing the detector for increased sample throughput. The on-line sample extraction step takes 15 seconds followed by gradient chromatography taking another 90 seconds. Having minimal sample pretreatment the method is both simple and fast. This system has been used to successfully develop a validated positive-ion electrospray bioanalytical method for the quantitation of vancomycin. Detection of vancomycin was accurate and precise, with a limit of detection of 1 ng/mL in serum and urine. The calibration curves for vancomycin in rat, dog and primate were linear in a concentration range of 0.001-10 microg/mL for serum and urine. This method has been successfully applied to determine the concentration of vancomycin in rat, dog and primate serum and urine samples from pharmacokinetic and urinary excretion studies.

Animals↗
PubMed2004

Ultraviolet matrix-assisted laser desorption/ionization time-of-flight mass spectrometry of intact hemoglobin complex from whole human blood.

Ultraviolet matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (UV-MALDI-TOFMS) was applied to the analysis of intact human hemoglobin complex directly from whole human blood. The less acidic matrix substance 2,6-dihydroxyacetophenone provided sufficient insensitivity to the impurities present in this crude biological matrix to make any sample pretreatment except dilution dispensable. This matrix facilitated exact molecular mass determination of the non-assembled hemoglobin alpha- and beta-chain (SD < or = +/-0.28 Da and Deltam < or =6.4 ppm for n = 10) if trifluoroacetic acid was used as an additive. Replacement of the denaturing additive trifluoroacetic acid by the neutral salt ammonium acetate allowed the detection of the intact hemoglobin alpha(2)beta(2)H(n)-assembly (n = 0-4) and the alphabeta-subassembly. A prerequisite for the detection of the noncovalent complex was the application of a very soft sample preparation procedure. Crystal morphology, sample concentration and laser pulse energy were also found to be important parameters for the analysis of the intact protein complexes. However, comparison with published electrospray ionization (ESI)-MS results on mammalian hemoglobin molecules shows that, even under the applied gentle conditions, MALDI does not provide a completely reliable picture of the solution-phase equilibrium. In contrast to ESI, especially extensive loss of the heme b (H) groups was noticed. The disruption of the rather stable heme b-globin interaction is assumed to be induced by photo-excitation during the desorption/ionization process.

Hemoglobins↗
PubMed1999

Quantitative determination of efavirenz (DMP 266), a novel non-nucleoside reverse transcriptase inhibitor, in human plasma using isocratic reversed-phase high-performance liquid chromatography with ultraviolet detection.

Efavirenz is a novel non-nucleoside reverse transcriptase inhibitor for the treatment of HIV-1-infected individuals. A simple and rapid high-performance liquid chromatographic method for the quantification of efavirenz in human plasma suitable for therapeutic drug monitoring in plasma is described. Sample pretreatment consists of protein precipitation with acetonitrile and subsequent evaporation of the extract to concentrate the analyte. The drug is separated from endogenous compounds by isocratic reversed-phase high-performance liquid chromatography with ultraviolet detection at 246 nm. The method has been validated over the range of 10 to 10,000 ng/ml using a volume of 250 microl of plasma. The assay is linear over this concentration range as indicated by the F-test for lack of fit. Within- and between-day precisions are less than 4.3% for all quality control samples. The lower limit of quantitation is 10 ng/ml and the recovery of efavirenz from human plasma is 106.4% (+/- 1.8%). Frequently co-administered drugs did not interfere with the described methodology. Efavirenz is stable under various relevant storage conditions, for example when stored for 24 h at room temperature. This validated assay is suited for use in pharmacokinetic studies with efavirenz and can readily be implemented in the setting of a hospital laboratory for the monitoring of efavirenz concentrations.

Acetonitriles↗
PubMed1999

Quantitative determination of abacavir (1592U89), a novel nucleoside reverse transcriptase inhibitor, in human plasma using isocratic reversed-phase high-performance liquid chromatography with ultraviolet detection.

Abacavir is a novel nucleoside reverse transcriptase inhibitor for the treatment of HIV-1 infection. A simple and rapid high-performance liquid chromatographic method for the quantification of abacavir in human plasma suitable for pharmacokinetic research purposes is described. Sample pretreatment consists of protein precipitation with perchloric acid. The supernatant is injected directly into the chromatographic system after centrifugation. The drug is separated from endogenous compounds by isocratic reversed-phase high-performance liquid chromatography with ultraviolet detection at 285 nm. The method has been validated over the range of 20-2000 ng/ml using a volume of 300 microl of plasma. The assay is linear over this concentration range as indicated by the F-test for lack-of-fit. Within- and between-day precisions are less than 7.5% for all quality control samples. The lower limit of quantitation is 20 ng/ml and the recovery of abacavir is 88.1% (+/-1.3%). Frequently coadministered drugs did not interfere with the described methodology. Abacavir is stable in human plasma under various relevant storage conditions, for example when stored for 51 days at -20 degrees C. This validated assay is suited for use in pharmacokinetic studies with abacavir in human plasma and can readily be implemented in the setting of a hospital laboratory for the monitoring of abacavir concentrations.

Acquired Immunodeficiency Syndrome↗