Identification and quantitation of gamma-hydroxybutyrate (NaGHB) by nuclear magnetic resonance spectroscopy

The most common means of identification of gamma-hydroxybutyrate (NaGHB) involves using Fourier transform infrared spectroscopy (FTIR) or gas chromatography-mass spectrometry (GC-MS) of a suitable derivative. However, these methods may be complicated by possible shifts in chemical equilibrium between gamma-hydroxybutyric acid (GHB), GHB salts and the precursor lactone, gamma-butyrolactone (GBL). This paper addresses the technique of proton and carbon nuclear magnetic resonance spectroscopy (1H and 13C NMR) for the direct and accurate identification of GHB and GBL. The application of 1H NMR for GHB quantitation is also discussed.     

Lifestyle illicit drug seizures: A routine ESI-LC-MS method for the identification of sildenafil and vardenafil

For street samples suspected of containing the phosphodiesterase-5 inhibitors sildenafil (Viagra(?)) and/or vardenafil (Levitra(?)), including powders or adulterated herbal supplements, a chemical analysis is needed to provide confirmatory identification of these illegally procured substances. Sildenafil and vardenafil are structurally similar and it is difficult to differentiate between them, as previous mass spectrometric studies have shown the two drugs to produce similar fragmentation patterns. The use of tandem mass spectrometry can produce confirmatory data, but the technique requires a high level of technical expertise. We have developed an electrospray ionization-liquid chromatography-mass spectrometry (ESI-LC-MS) method that allows differentiation between these two structurally similar molecules via in-source fragmentation in combination with an ion trap mass spectrometer. A very stable gas phase ion is formed during in-source fragmentation of vardenafil; the combination of the stability of this ion and the longer residence time for the ion in the ion trap results in a very strong signal. This feature results in a method that can provide clear differentiation between sildenafil and vardenafil while at the same time requiring less expertise from the routine analyst to confirm the presence or absence of the two compounds.     

A methodology based on NMR spectroscopy for the forensic analysis of condoms.

Both solution and solid state Nuclear Magnetic Resonance (NMR) spectroscopic techniques have been used to determine differences in commercially available condoms. Whilst solid state NMR is useful for determining the polymer backbone, it is not useful for forensic analysis due to the commonality of the latex condom. However solution NMR spectra obtained following a simple extraction procedure using hexane, provides a fingerprint of the additives in the lubricants. Following the development of a flow chart, basing decisions on the presence of particular peaks present in the solution spectra, 33 of 38 condoms could be individualized. Samples were also analyzed after having the lubricant manually removed and soaking the condom in water for 3 to 24 h. These experiments were performed to simulate a case of the sample having been used and disposed of by flushing down the toilet, as may be experienced in a case of a sexual assault. The results indicated that the only significant water soluble component was polyethylene glycol. The overall results suggest that the method developed may be a quick and useful technique in characterizing condoms. The information obtained can be used to provide associative evidence between suspect and crime, and so be useful in sexual assault cases.     

Measuring the thickness of facial soft tissues using nuclear magnetic resonance tomography for the purpose of identification

Nuclear spin tomography since its beginnings in the seventies has steadily gained in importance as a method of examination in medical diagnostics as it produces a picture. In the field of forensic medicine the NMR technique as used for anatomic-anthropologic issues attempting to identify skulls this is a valuable supplement to an extension of the existing methods of investigation. The results of a measurement of the thickness of soft facial tissue in a live test person is shown as compared to measures obtained by sonography.     

Virtopsy,a new imaging horizon in forensic pathology: virtual autopsy by postmortem multislice computed tomography (MSCT) and magnetic resonance imaging (MRI)--a feasibility study

Using postmortem multislice computed tomography (MSCT) and magnetic resonance imaging (MRI), 40 forensic cases were examined and findings were verified by subsequent autopsy. Results were classified as follows: (I) cause of death, (II) relevant traumatological and pathological findings, (III) vital reactions, (IV) reconstruction of injuries, (V) visualization. In these 40 forensic cases, 47 partly combined causes of death were diagnosed at autopsy, 26 (55%) causes of death were found independently using only radiological image data. Radiology was superior to autopsy in revealing certain cases of cranial, skeletal, or tissue trauma. Some forensic vital reactions were diagnosed equally well or better using MSCT/MRI. Radiological imaging techniques are particularly beneficial for reconstruction and visualization of forensic cases, including the opportunity to use the data for expert witness reports, teaching, quality control, and telemedical consultation. These preliminary results, based on the concept of "virtopsy," are promising enough to introduce and evaluate these radiological techniques in forensic medicine.     

The effect of exposure to the elements on the forensic characterization by infrared spectroscopy of poly(ethylene terephthalate) fibers

Validation of a recently proposed differentiation method for single poly (ethylene terephthalate) (PET) fibers, based on integration of IR absorption bands, is presented. IR microscopy in the transmission mode, differential scanning calorimetry and optical microscopy were employed. The reliability of the technique has been extended to the cases in which the fibers being compared have experienced long exposure to the elements, as may happen if these items are recovered from open-air crime scenes. The effect of immersion in water and irradiation by the sun for a three-month period have been considered. The fibers remained morphologically and structurally unaltered. The significance of comparisons between fibers kept in different environments is thus enhanced, because the detrimental effect of the elements can be ruled out. This further validation of the proposed technique contributes to render the data presented by the forensic scientist less vulnerable to context effect and bias.     

Application of dispersive liquid-liquid microextraction and CE with UV detection for the chiral separation and determination of the multiple illicit drugs on forensic samples

A new method was developed for pre-concentration, chiral separation and determination of multiple illicit drugs on forensic samples using dispersive liquid-liquid microextraction (DLLME) and capillary electrophoresis (CE) with Ultra Violet (UV) detection. The method was based on the formation of tiny droplets of an organic extractant in the prepared sample solution using water-immiscible organic solvent (chloroform) dissolved in water-miscible organic dispersive solvent (isopropyl alcohol). The organic phase, which extracted heroin, DL-methamphetamine, DL-3, 4-methylenedioxymethamphetamine and dl-ketamine from the prepared sample solution, was separated by centrifuging. The sedimented phase was transferred into a small volume CE auto-sampler vial with 10 μL of 1% HCl methanol solution and evaporated to dryness. The residue was reconstituted in lidocaine hydrochloride aqueous solution (internal standard) and introduced by electrokinetic injection into CE. Parameters affecting extraction efficiency were investigated and optimized. Under optimum conditions, linearity of the method was 0.15-6500 μg/L for all target analytes. The LODs (S/N=3) were 0.05-0.20 μg/L. Excellent repeatability (RSD ≤ 4.4%, n=5) was achieved. The feasibility of this method was demonstrated by analyzing spiked forensic samples. To our knowledge, it is the first time to combine DLLME with CE for chiral separation and determining illicit drugs on forensic samples.     

Determination of paraquat in raw and formalin-fixed tissues by electron spin resonance (ESR) spectroscopy

ESR method was applied to determine paraquat levels in fresh and formalin-fixed tissues. Paraquat was converted to paraquat radical by adding sodium dithionite to tissue homogenates and detected by ESR. Paraquat levels of more than 0.2 micrograms/ml homogenate could be quantified with 0.1 ml of the homogenate. The use of manganese ions for standardization of paraquat signal enabled much more accurate ESR measurements because this ion was quite stable and its signal did not overlap that of paraquat. Even with tissues fixed in formalin, tissues paraquat levels were measureable after removing formalin from the tissue extract. This fact was verified by studying two cases; the tissues were kept in formalin for 1.5 years in case 1 and for 6.5 years in case 2. In both cases, the paraquat contents in tissues were 0.02-0.08 micrograms/g. In this way ESR is one of the most suitable methods in determining low levels of paraquat in tissues even after they were preserved in formalin for a long time.     

The forensic chemical analysis of cadaveric material for the presence of drug and narcotic compounds]

The authors consider that the list of drugs and narcotics which can be reliably identified in a general forensic chemical analysis of cadaveric material (not a purposeful screening) should be changed. Isolation by acidified water, hydrochloric acid hydrolysis, and by acetonitrile should be used for isolation of drugs and narcotic compounds in forensic chemical analysis of cadaveric material.     

Drugs WorkBook (DWB): A tool for the analysis of illicit drugs in seized materials

Accurate and reliable analytical measurements are essential when data are to be used to assist the Court in deciding whether or not a drug offence has been committed and therefore about either the innocence or guilt of the accused. The Italian law on drugs demands that compliance with specification limits be assessed on the basis of the actual content of controlled substance contained in seized materials. As a consequence, the role of measurement uncertainty, significant figures and rounding errors becomes critical. In order to assist analysts of forensic toxicology laboratories with illicit drug-related cases, a software tool named Drugs WorkBook (DWB) has been developed. The tool is useful for the quantification of illicit drugs in seized materials along with their measurement uncertainties, the assessment of compliance to specification limits, the printing of comprehensive laboratory reports and the organization of case archives. Other quality control topics, such as control charts, are included. The tool's databases can be edited by the user and maintained up to date. The tool is made freely available to the scientific community.     

mRNA在体液斑鉴定研究中内参基因的选择与应用

利用mRNA分析技术鉴定生物检材中各种体液斑迹的组织属性来源是近年来法医物证学的一个重要进展。终点逆转录PCR和实时荧光定量PCR技术是法医学实验室常用的mRNA检测技术,为保证实验结果的准确性,研究者通常选用组成性表达的管家基因作为内参,对不同样本提取RNA进行质控和标准化,以正确评估目标mRNA在各样本中的表达量。本文聚焦近年来使用mRNA分析技术进行体液斑迹组织属性来源鉴定的研究报道,对这些工作中内参基因的选择与应用效果进行综述。     

Current status and prospects for developing chemical-toxicological (forensic chemical) studies in the Russian Federation

The authors discuss modern state and prospects of planning and carrying out research in forensic (toxicological) chemistry within the framework of practical forensic medical expert evaluation, state of forensic medical expert evaluation of alcohol and narcotic intoxication, and other pressing problems of forensic medical expert practice.     

Sensitive and specific multiresidue methods for the determination of pesticides of various classes in clinical and forensic toxicology

Original and sensitive multiresidue methods are presented for the detection and quantitation, in human biological matrices, of 61 pesticides of toxicological significance in human. These methods involved rapid solid-phase extraction using new polymeric support (HLB and MCX) OASIS cartridges. Gas chromatography-mass spectrometry (GC-MS) was used for volatile (organophosphate, organochlorine, phtalimide, uracil) pesticides and liquid chromatography-ionspray-mass spectrometry (LC-MS) for thermolabile and polar pesticides (carbamates, benzimidazoles). Acquisition was performed in the selected ion monitoring (SIM) mode. Extraction recovery varied owing to the nature of pesticides, but was satisfactory for all. Limits of detection (LODs) and limits of quantitation (LOQs) ranged, respectively, from 2.5 to 20 and from 5 to 50ng/ml. An excellent linearity was observed from LOQs up to 1000ng/ml for all the pesticides studied. The proposed procedures yielded reproducible results with good inter-assay accuracy and precision. A few cases of intoxication are presented to demonstrate the diagnostic interest of these methods: in two cases were determined lethal concentrations of endosulfan and carbofuran; in four other cases, the procedures helped diagnose intoxication with, respectively, parathion-ethyl, the association of bromacil and strychnine, bifenthrin and aldicarb.     

Developmental validation of a multiplex qPCR assay for assessing the quantity and quality of nuclear DNA in forensic samples

Forensic scientists are constantly searching for better, faster, and less expensive ways to increase the first-pass success rate of forensic sample analysis. Technological advances continue to increase the sensitivity of analysis methods to enable genotyping of samples containing minimal amounts of DNA, yet few tools are available that can simultaneously alert the analyst to both the presence of inhibition and level of degradation in samples prior to genotyping to allow analysts the opportunity to make appropriate modifications to their protocols and, consequently, to use less sample. Our laboratory developed a multiplex quantitative PCR assay that amplifies two human nuclear DNA target sequences of different length to assess DNA degradation and a third amplification target, a synthetic oligonucleotide internal PCR control (IPC), to allow for the assessment of PCR inhibition. We chose the two nuclear targets to provide quantity and fragment-length information relevant to the STR amplification targets commonly used for forensic genotyping. The long target (nuTH01, 170-190 bp) spans the TH01 STR locus and uses a FAM-labeled TaqMan probe for detection. The short nuclear target (nuCSF, 67 bp) is directed at the upstream flanking region of the CSF1PO STR locus and is detected using a VIC-labeled TaqManMGB probe. The IPC target sequence is detected using a NED-labeled TaqManMGB probe. The assay was validated on the Applied Biosystems 7500 Real-Time PCR system, which is optimized for NED detection. We report the results of a developmental validation in which the assay was rigorously tested, in accordance with the current SWGDAM guidelines, for precision, sensitivity, accuracy, reproducibility, species specificity, and stability.