Automated identification of toxic substances in poisoned human fluids by a retention prediction system in reversed-phase liquid chromatography

An automated identification system based on the retention prediction concept has been constructed for toxic substances. The system performance has been evaluated for the identification of toxic compounds in poisoned human fluids.     

Microcomputer-assisted liquid chromatographic separation system: application to toxic compounds identification in poisoned human fluids

The computer-assisted liquid chromatographic system (MCASYST) is developed for automated identification and analysis. The system has six main functions: retention prediction system, liquid chromatographic data base system, automated identification system, optimization of separation conditions, data loading program from UV multichannel detector, and UV spectral data base system. The performance and potential of this MCASYST system is evaluated for toxic compounds identification in poisoned human fluids.     

Utilization of Automatically Assigned Retention Indices for Computer Identification of Mass Spectra

Abstract

A computer program which provides retention indices for the entire gas chromatogram has been applied to assist in the identification of components of mixtures using a GC-MS-Computer system. As one example, the identification of drugs and metabolites in body fluids is discussed and it is shown that retention indices, in combination with mass spectral data, allow a most efficient, unambiguous and readily automated identification of components of the relatively complex mixtures which result from the extraction of body fluids.     

导电球简立方格子有效介电常数的计算

Based on the Maxwell-Wagner model, an analytical formula for effective dielectric constants is derived as a series expansion in powers of the volume fraction of spheres. Effective dielectric constants of simple cubic lattices of conducting particles suspended in dielectric or conducting fluids are calculated. The numerical results show that effective dielectric constants depend upon the ratios of the permeability of conducting spheres to that of the suspending fluids under high frequency (0.1-1 kHz) applied fields, whereas, it is determined by the ratios of the conductivities of spheres to that of fluids under low frequency or dc electric fields. The imaginary parts of effective dielectric constants can be very big sometimes. This means that the resistive losses of electrorheological fluids can be very strong at times. The effect of conduction in a system cannot be neglected in the design of high performance electrorheological fluids.     

Separation of thiazide diuretics and antihypertensive drugs by thin-layer chromatography.

A procedure is reported for the thin-layer chromatographic (TLC) separation and identification of thiazide diuretics and other antihypertensive drugs. Several new solvent systems and a variety of possible detection reagents were examined. Twenty thiazides used routinely in therapy were successfully separated and identified. Use of the TLC systems and combinations of the detecting methods described should be useful for the identification of these drugs in biological fluids and in various dosage forms.     

Extended specificity studies of mRNA assays used to infer human organ tissues and body fluids

Messenger RNA (mRNA) profiling is a technique increasingly applied for the forensic identification of body fluids and skin. More recently, an mRNA‐based organ typing assay was developed which allows for the inference of brain, lung, liver, skeletal muscle, heart, kidney, and skin tissue. When applying this organ typing system in forensic casework for the presence of animal, rather than human, tissue is an alternative scenario to be proposed, for instance that bullets carry cell material from a hunting event. Even though mRNA profiling systems are commonly in silico designed to be primate specific, physical testing against other animal species is generally limited. In this study, human specificity of the organ tissue inferring system was assessed against organ tissue RNAs of various animals. Results confirm human specificity of the system, especially when utilizing interpretation rules considering multiple markers per cell type. Besides, we cross‐tested our organ and body fluid mRNA assays against the target types covered by the other assay. Marker expression in the nontarget organ tissues and body fluids was observed to a limited extent, which emphasizes the importance of involving the case‐specific context of the forensic samples in deciding which mRNA profiling assay to use and when for interpreting results.     

Rapid diagnosis of drug intoxication using novel NAGINATA gas chromatography/mass spectrometry software

In Japan, not only the classical stimulant, methamphetamine, but also a wide variety of illicit drugs and designer drugs are abused by juveniles. It is, however, difficult to screen these drugs in human urine due to the poor availability of high-quality standards. Therefore, it is important to develop a screening method that does not require the use of standard compounds. Furthermore, if we can obtain approximate drug concentrations in biological fluids by the first screening procedure, the subsequent treatment of the patient and forensic diagnosis can be carried out more rapidly and exact quantitative analysis performed more efficiently. We have devised a rapid screening method for the simultaneous semi-quantitative analysis of 30 abused drugs using gas chromatography/mass spectrometry (GC/MS) with a retention time locking technique. Based on this method, an 'abused drugs database' was constructed including retention time (RT), qualifier ion/target ion (QT) percentage and calibration curve (values of slope and intercept) using the novel GC/MS software, NAGINATA. We compared the analytical results obtained by this method using the constructed database with those from conventional methods in six forensic cases. The number of confirmed drugs and concentrations obtained by the established method was comparable with that obtained by conventional methods. We found a significant improvement in the time for data analysis, and qualitative and quantitative information about each drug was obtained without using standards. Therefore, this new screening procedure using NAGINATA has potential for the rapid identification of poisoning and should be useful in clinical and forensic toxicological analyses.     

The potential of serially coupled alkyl-bonded silica monolithic columns for high resolution separations of pharmaceutical compounds in biological fluids

Summary In this paper we investigate the potential of alkyl-bonded silica monolithic columns for the isolation and identification of drug-related components in biological fluids. Up to 6 columns have been connected in series to produce a chromatographic system with up to 40,000 plates. This high-resolution chromatography system has been coupled to both MS and NMR to enable efficient detection and characterisation of drug-related components in biological fluids. The use of six coupled columns has been shown to give enhanced resolution over a high quality silica particulate column packed with 3 μm material which exhibits the same back pressure. The effect of volume and mass load on the performance of monolithic columns for semi-preparative chromatography of biological fluids has also been investigated. In these studies it was possible to inject up to 100 mL of neat urine with no loss of chromatographic performance. Furthermore, upon re-testing, the columns showed similar chromatographic performance. Again several columns were serially connected, producing enhanced resolution in the semi-preparative mode.     

Practical Microbore Column HPLC: System Development and Drug Applications

Abstract

Commercially available packed microbore columns have been used to demonstrate the potential of microbore high performance liquid chromatography (micro-LC) systems. The necessity of optimizing the chromatographic system for micro-LC is demonstrated using sulfa drugs and antibiotic standards, and some of the advantages of micro-LC such as high mass sensitivity are shown. Finally micro-LC has been used for the determination of two drugs, reserpine and trichlormethiazide in biological fluids.     

Chromatographic screening for drugs of abuse using capillary columns II. Peak identification on support coated open tubular columns for a series of central nervous system stimulant drugs

Summary A system based on Kovats' Retention Indices is described for the identification of CNS stimulant drugs recovered from body fluids. Use is made of the difference in retention indices found on polar and non-polar GLC columns (l values) and the effect of operating temperature is discussed.     

Identification of novel circulating coffee metabolites in human plasma by liquid chromatography-mass spectrometry

This study reports a liquid chromatography-mass spectrometry method for the detection of polyphenol-derived metabolites in human plasma without enzymatic treatment after coffee consumption. Separation of available standards was achieved by reversed-phase ultra performance liquid chromatography and detection was performed by high resolution mass spectrometry in negative electrospray ionization mode. This analytical method was then applied for the identification and relative quantification of circulating coffee metabolites. A total of 34 coffee metabolites (mainly reduced, sulfated and methylated forms of caffeic acid, coumaric acid, caffeoylquinic acid and caffeoylquinic acid lactone) were identified based on mass accuracy (<4 ppm for most metabolites), specific fragmentation pattern and co-chromatography (when standard available). Among them, 19 circulating coffee metabolites were identified for the first time in human plasma such as feruloylquinic acid lactone, sulfated and glucuronidated forms of feruloylquinic acid lactone and sulfated forms of coumaric acid. Phenolic acid derivatives such as dihydroferulic acid, dihydroferulic acid 4'-O-sulfate, caffeic acid 3'-O-sulfate, dimethoxycinnamic acid, dihydrocaffeic acid and coumaric acid O-sulfate appeared to be the main metabolites circulating in human plasma after coffee consumption. The described method is a sensitive and reliable approach for the identification of coffee metabolites in biological fluids. In future, this analytical method will give more confidence in compound identification to provide a more comprehensive assessment of coffee polyphenol bioavailability studies in humans.     

Separation and identification of sugars and maltodextrines by thin layer chromatography: Application to biological fluids and human milk

A monodimensional thin layer chromatography method to separate several sugars of clinical interest is described. The separation and identification of 14 sugars (L-fucose, D-galactose, D-glucose, lactose, N-acetylglucosamine, D-maltose, D-manose, L-sorbase, fructose, D-xylose, glucuronic acid, N-acetyllactosamine, 3' and 6' sialyllactose) and maltodextrines (G(2)-G(8)) is possible by using two different eluents mixtures, as well as two different detection reagents. The method has been applied to separate sugars, maltodextrines and oligosaccharides in several biological fluids (blood, urine and faeces), in an infant milk and in human milk. It is a very simple technique (with a high sensitivity) that can be used in any lab.     

High-performance liquid chromatographic determination of chloramphenicol and four analogues using reductive and oxidative electrochemical and ultraviolet detection

A high-performance liquid chromatographic procedure is described for the separation, quantitation and identification of chloramphenicol, dehydrochloramphenicol, nitrophenylaminopropanedione, nitrosochloramphenicol and aminochloramphenicol. An isocratic reversed-phase system with ultraviolet and electrochemical detectors in tandem was assembled and used. The system was constructed with special accommodation to enable us to use the electrochemical detector in both reductive and oxidative modes. Retention characteristics, hydrodynamic voltammograms under reductive and oxidative conditions and ultraviolet absorbance are reported. Applicability of the procedure to biological fluids was demonstrated by separation and detection of chloramphenicol after incubation with human blood.     

Electromembrane extraction from biological fluids

Electro-assisted extraction of ionic drugs from biological fluids through a supported liquid membrane (SLM) and into an aqueous acceptor solution was recently introduced as a new sample preparation technique termed electromembrane extraction (EME). The applied electrical potential across the SLM has typically been in the range of 1-300 V. Successful extractions have been demonstrated even with common batteries (9 V) instead of a power supply. The chemical composition of the SLM has been crucial for the selectivity and for the recoveries of the extraction. Compared to other liquid-phase microextraction techniques (LPME), extraction times have been reduced by a factor of 6-17, and successful extractions have been obtained at extraction times of 1-5 min, and even down to a few seconds with online microfluidic EME devices. The technique has provided very efficient sample clean-up and has been found well suited for the extraction of sample sizes in the low μL range. Extractions have been performed with both rod-shaped hydrophobic porous fibers and with flat hydrophobic porous sheets as SLM support. The technique has been successfully downscaled into the micro-chip format. The nature of the SLM has been tuned for extraction of drugs with different polarity allowing extractions to be tailored for specific applications depending on the analyte of interest. The technique has been found to be compatible with a wide range of biological fluids and extraction of drugs directly from untreated human plasma and whole blood has been demonstrated. EME selectively extracts the compounds from the complex biological sample matrix as well as allowing concentration of the drugs. With home-built equipment fully acceptable validation results have been obtained.     

Identification of the major metabolites of (R)‐salbutamol in human urine,plasma and feces using ultra high performance liquid chromatography coupled with quadrupole time‐of‐flight mass spectrometry

(R)‐Salbutamol is a selective β2‐adrenoreceptor agonist, which produces a short‐acting bronchodilator effect and is widely used for the treatment of respiratory diseases in humans. Drug metabolism and identification of the metabolites play an essential role in the evaluation of the overall efficacy and safety of the drugs in clinical practices. There are few reports on the identification of major metabolites of (R)‐salbutamol in humans, and the number of identified metabolites is very limited. In this research, a method of ultra‐high performance liquid chromatography coupled with quadrupole time‐of‐flight mass spectrometry was developed for the discovery and identification of (R)‐salbutamol and its major metabolites in human biological samples. Totally, twelve metabolites of (R)‐salbutamol were found and identified and all the metabolites could be found in urine, one metabolite in plasma and two metabolites in feces. Among all the metabolites, eight metabolites have never been reported before. The results indicated that (R)‐salbutamol was mainly metabolized through isomerization, oxidation, reduction, glucuronidation, and sulfation pathways in vivo. The possible metabolic pathways of (R)‐salbutamol were subsequently presented in this study, which contribute to a better understanding of the metabolism of (R)‐salbutamol in humans.     

Microneedles-based electrochemical sensors: New tools for advanced biosensing

Electrochemical biosensors are used worldwide as analytical tools from laboratory applications to market products. The performance of electrochemical sensing can be boosted by adopting the microneedle (MN) geometry as an innovative configuration of standard electrodes. MNs can be miniaturized, easily functionalized, and properly designed for specific aim monitoring, but most of all, they allow a low invasive controlling tool for growth and for environment influence in plant and a painless door to human body fluids where target analytes can be detected, overcoming the natural barrier of the skin. In this review, the very recent developments in MN-based electrochemical biosensing published in the literature are summarized.     

Proteomic profiling of human urine using multi-dimensional protein identification technology

Human urine samples are ideal for proteomic profiling and have tremendous potential as sources of biomarkers. Multi-dimensional protein identification technology (MudPIT) is an effective approach to analyzing human urine or other fluids dominated by diverse metabolites. MudPIT analysis was used to identify 87 proteins in just 15 ml of human urine. A high throughput, reproducible, and sensitive technology, MudPIT may soon be used for more proteomic analyses of metabolites.     

On-line diode array UV-visible spectrometry in screening for drugs and drug metabolites by high-performance liquid chromatography

The direct coupling of a multi-channel diode array UV-visible spectrophotometer to a powerful reversed-phase HPLC separation system is considered, especially for use in qualitative analysis, e.g., screening/identification of drugs and drug metabolites. The approach is illustrated by the screening for metabolites of butoprozine and ticlopidine directly in human and rat bile.     

The use of the PREP system for handling of body fluids

The Du Pont PREP automated sample processor is a centrifugally based, microprocessor controlled instrument that was designed for extraction of samples from biological fluids. Extraction takes place in cartridges containing either organic resins or bonded silica packings as extraction sorbants. This paper will discuss the application of several lipophilic and ion exchange sorbants to the extraction of biological samples from body fluids. The advantages of these different types of sorbants will be compared and their performance with automated sample preparation will be shown. A variety of applications including the extraction of benzodiazepine, barbiturate, aminoglycoside and anticonvulsant drugs and their metabolites from serum, urine, and tissue homogenates will be discussed.     

Selective system of identification and determination of antidepressants and neuroleptics in serum or plasma by solid-phase extraction followed by high-performance liquid chromatography with photodiode-array detection in analytical toxicology.

A selective off-line solid-phase isolation of antidepressants, neuroleptics and other structurally related basic drugs from plasma or serum prior to high-performance liquid chromatography was tested and optimized for general use in toxicological analyses where concomitant drugs can be encountered. The sequential elution preseparated drug mixtures and simplified the subsequent analytical steps. High isolation efficiencies of cyano-bonded silica cartridges from Baker for fifteen amine drugs were determined. Isocratic chromatography on octadecylsilica proved to be very suitable for broad practical applications in complicated cases. The identification of an unknown peak was supported by photodiode-array detection in the range 200-400 nm with a resolution of 2 nm. The linearity of the assay from therapeutic to toxic concentrations was attained. Sufficient sensitivities covering low therapeutic levels of parent drugs and their demethylated metabolites were reached. The system is flexible and allows various methods of quantitative assay to be devised according to the conditions of a particular case in clinical or forensic toxicology.