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G Rule

Publications and source records attributed to G Rule.

6 recordsLinked to original sources
PubMed2001

A 384-well solid-phase extraction for LC/MS/MS determination of methotrexate and its 7-hydroxy metabolite in human urine and plasma.

A solid-phase extraction procedure, in a 384-well format, has been developed for methotrexate and its primary metabolite, 7-hydroxymethotrexate, in human urine and plasma. This format has not been utilized previously for solid-phase extraction of drugs from biological fluids. The 384-well plates contained a C-18 stationary phase bonded to silica particles which are incorporated into a glass-fiber membrane. Methotrexate and 7-hydroxymethotrexate have been quantified across the curve range of 1 to 50 microg/mL and 50 to 1000 ng/mL, respectively, in urine and from 5 to 250 ng/mL and 5 to 100 ng/mL, respectively, in plasma. Both analytes are quantified by linear regression using 20-microL sample aliquots. Experiments to evaluate the influence of particle size, elution volume, and injection volume on signal intensity were conducted and are reported, along with the results of experiments examining cross contamination between wells. Recovery was determined to be > or = 95% from urine. Results from a run of 384 samples analyzed over a 14-h period indicate that 384-well SPE can be successfully utilized to increase analytical run sizes and sample throughput for LC/MS/MS determination of small drug molecules in biological samples.

Chromatography, Liquid↗
PubMed1999

High-throughput sample preparation and analysis using 96-well membrane solid-phase extraction and liquid chromatography-tandem mass spectrometry for the determination of steroids in human urine.

A 96-well solid-phase extraction (SPE) system is used to rapidly prepare human urine samples for high-throughput quantitative analysis of two steroids, equilenin and progesterone, by liquid chromatography-tandem mass spectrometry using deuterated estrone as the internal standard. We define high-throughput here as analysis of 384 samples in a 24 h period. A total of 384 samples and standards were extracted by an individual in one day and subsequently analyzed within a 24 h period. The inter- and intratray accuracy and precision obtained over the course of these injections was within 8% coefficient of variation when analyzed by atmospheric pressure chemical ionization mass spectrometry using positive ion detection. A semiautomated sample processing workstation was used to add internal standard and then process 96 samples at a time. The recovery of the analytes from the SPE was approximately 85%. The accuracy and precision obtained was comparable to that ordinarily obtained using manual sample preparation techniques.

Autoanalysis↗
PubMed1993

A quest for oleandrin in decayed human tissue.

This forensic case taught us several lessons. First, there is a need for improved sample cleanup and treatment of severely decayed tissue samples when trace determinations of target analytes are needed. With the exception of a few reports the literature is lacking in information with regard to the most modern sample preparation techniques. Second, the coupling of LC/LC with tandem MS provides an effective means of "on-line" samples cleanup for complex sample matrices. The improvements in selectivity shown in Figure 3 reveal the analytical power available when these techniques are combined. Third, once we decided to use LC/LC/MS/MS, we were able to analyze more than 50 samples in a semi-automated fashion over approximately three days. The reliability and ruggedness of the combined techniques and equipment suggest this approach may have merit for common applications in which large numbers of biological samples (e.g., plasma and urine) must be analyzed. As a postscript, when this project was completed we proposed that the use of antibodies for isolating oleandrin and its relatives might be a more selective means for trace enrichment of the target analytes. For example, a high-pressure immunoaffinity column could have been coupled on line as column 1 in Figure 4. After pumping a relatively high volume of aqueous tissue extract through an immunoaffinity column during trapping and trace enrichment conditions, the column could be rinsed with phosphate-buffered saline. Then the pH could be lowered to unfold the antibody protein and allow release of the trapped analyte from this column with subsequent trapping on column 2 in Figure 4.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1981

Liposome-mediated delivery of deoxyribonucleic acid to cells: enhanced efficiency of delivery related to lipid composition and incubation conditions.

Delivery of liposome-encapsulated simian virus 40 (SV40) DNA to African green monkey Related to been used as a probe to study liposome--cell interactions and to determine conditions which favor the intracellular delivery of liposome contents to cells. The efficiency of DNA delivery by various liposome preparations (monitored by infectivity assays) was found to be dependent both on the magnitude of vesicle binding to cells and on the resistance of liposomes to cell-induced leakage of contents. Acidic phospholipids were much more effective in both binding and delivery, and phosphatidylserine (PS) was the best in both aspects. The inclusion of 50 mol % cholesterol in liposomes reduces the cell-induced leakage of vesicle contents (2--5-fold) and substantially enhances the delivery of DNA to cells (2--10-fold). Following incubation of cells with negatively charged liposomes containing SV40 DNA, infectivity can be enhanced greatly by brief exposure of the cells to glycerol solutions. In contrast, only slight enhancement by glycerol was observed for SV40 DNA encapsulated in neutral or positively charged liposomes. The results of competition experiments between empty phosphatidylcholine liposomes and DNA-containing PS liposomes also suggest possible differences in the interaction of neutral and negatively charged liposome preparations with cells. Morphological studies indicate that the glycerol treatment stimulates membrane ruffling and vacuolization and suggest that the enhanced uptake of liposomes occurs by an endocytosis-like process. Results obtained with metabolic inhibitors are also consistent with the interpretation that the enhancement of liposome delivery in glycerol-treated cells occurs via an energy-dependent endocytotic pathway. Pretreatment of cells with chloroquine, a drug which alters lysosomal activity, further enhanced infectivity in glycerol-treated cells (4-fold). This observation suggests the involvement of a lysosomal processing step at some point in the expression of liposome-encapsulated DNA and, more importantly, illustrates the possibility of altering cellular mechanism to engineer more efficient delivery by liposomes. Under optimal conditions determined in this study, the efficiency of liposome-mediated SV40 DNA delivery was increased more than 1000-fold over that obtained by simply incubating cells with liposomes. It is also demonstrated that these conditions enhance delivery of other molecules, besides DNA, which are encapsulated in liposomes.

Animals↗
PubMed1979

Flow-injection analysis with chemiluminescence detection.

In "flow-injection analysis," the sample is inserted into a stream of reagent by use of a sample-injection valve. Mixing occurs downstream from the valve in a coil of tubing. With chemiluminescence detection this coil is positioned in front of a photomultiplier. We have evaluated the system for detection of hydrogen peroxide, using the luminol reaction with cupric ion as a catalyst. The effects of flow rate, sample volume, and reaction kinetics on the magnitude, duration, and repeatability of the chemiluminescent response have been evaluated. Precisions of 1 to 2% relative standard deviation on replicate measurements are readily achievable. A sample throughput of six samples per minute is possible with very little peak overlap. This detection system can be coupled to any process in which peroxide is generated.

Autoanalysis↗