New method of structural analysis and measurement of V-shaped percussion cracks in quartz sands surface by confocal laser scanning microscope (CLSM).

Microtextures of quartz sand grains can be used to establish sedimentary provenance of sedimentary deposits. V-shaped percussion cracks (Vs), which are randomly produced by grain-to-grain mechanical collision in high-energy subaqueous environments, are significant microtextures used for investigation of fluvial and marine sediment. In previous studies, others have used the percentage of microtextures obtained by scanning electron microscope to compare sediment samples. We developed a new method of measurement and evaluation of the surface of quartz grains using a confocal laser scanning microscope (CLSM) and applied the method to coastal sands containing quartz grains with abundant Vs. The method using international-standard surface-roughness parameters (ISO 25178) is useful for evaluating surface textures of quartz grains and for quantifying the structures of Vs (such as aperture area, density, and depth). The results of applying the method to coastal sands suggest that the density of Vs related to the coastal geological setting, that the depth of Vs related to wave height and offshore gradient, and that the size of Vs was not influenced by grain size or mineral composition of the coastal sands. This new method can corroborate preceding methods in provenance study of quartz grains in subaqueous environments. In principle, it also could apply to other types of microtextures and other minerals, such as degrees of weathering on surfaces of heavy minerals. This method, using a CLSM, has the potential to be applied to various provenance studies using grain-surface texture.

Analysis of water-soluble paper for forensic discrimination.

Water-soluble paper (WSP), which easily dissolves in water, has been found in criminal gang hideouts as evidence in crimes including bank transfer fraud, phone fraud, and grandparent scams, i.e. identity fraud to steal money. Distinguishing WSP products used in crimes is required in forensic investigations to link fraud groups with paper evidence or prove connections among fraud groups. In this work, we investigate distinguishing WSP products. White sheets of six WSP products available in Japan were analyzed by measuring the grammage and thickness, determining additive compounds by X-ray diffraction (XRD), X-ray fluorescence (XRF), and scanning electron microscopy (SEM)/energy dispersive X-ray spectrometry (EDX), and performing fiber analysis by transmitted light microscopy. The six products were categorized into three groups by grammage and thickness. XRD, XRF, and SEM/EDX analysis provided elemental information about additive compounds and their distribution on the surface. Pulp analysis by Graff "C" stain provided the composition and morphology of the pulp. The six products could be distinguished in a similar way to plain paper by these analyses. Our results demonstrate that conventional analytical methods used for plain paper analysis can also be used to discriminate WSP in forensic investigations.

Semi-automated scanning electron microscopy energy dispersive X-ray spectrometry forensic analysis of soil samples.

Soil is commonly analyzed to provide evidence because soil particles adhere to almost everything that may be of forensic interest. Particles derived from soil can be quantitatively analyzed by scanning electron microscopy with energy dispersive X-ray spectrometry (SEM-EDS). We developed a semi-automated SEM-EDS method for analyzing soil minerals. Soil was analyzed using the SEM-EDS automated particle analysis program, then the mineral species were identified from their chemical components. Chi-squared tests were used to discriminate between different minerals, and these tests were also applied to soil samples. The method put samples with the same parent population into the same group and discriminated between samples with different parent populations even when 26 particles were analyzed. The method successfully identified most soil samples with different parent populations as different. The results proved that the method could be used in forensic cases and provide new insights into the forensic analysis of soil samples.

Comparison and evaluation of the quick purification methods of methamphetamine hydrochloride from dimethyl sulfone for spectroscopic identification.

Methods to quickly purify methamphetamine hydrochloride from the cutting agent dimethyl sulfone for subsequent identification of confiscated crystalline samples using infrared absorption spectroscopy were compared and evaluated. Although sequential solvation and reprecipitation methods were simple, spectral contamination from dimethyl sulfone was inevitable and might affect the interpretation of the spectra. In addition, methamphetamine hydrochloride and dimethyl sulfone could form a solid solution because of solvation of both crystals into a single solution layer. By contrast, sublimation was an effective method for separation of methamphetamine hydrochloride and dimethyl sulfone. Sublimation combined with infrared absorption spectroscopy enabled rapid identification of crystalline methamphetamine hydrochloride.

Species identification of white false hellebore (Veratrum album subsp. oxysepalum) using real-time PCR.

Food poisoning is frequently caused by the accidental ingestion of toxic plants that possess strong morphological similarities to edible plants. False helleborine (Veratrum album) is one of the most common plants involved in such accidents. In cases of poisoning by toxic plants, rapid and accurate identification, usually based on the morphological or chemical analysis of plant parts, is required for appropriate medical treatment or forensic investigation. However, morphological examinations require experience in systematic botany because the samples are fragmentary, and chemical analysis of natural compounds can be difficult. In this study, we developed a TaqMan real-time PCR method using trnH-psbA and trnL-trnF that could be carried out in 30-60min. The lower detection limit was less than 10pg of DNA and the primer sets were specific to V. album and Veratrum stamineum. Mixed samples, cooked samples, and simulated gastric contents were successfully identified, and a multiplex assay of two regions was also possible. These results indicate that the TaqMan real-time PCR analysis is a very effective method to detect small samples of V. album and V. stamineum accurately and rapidly in poisoning cases.

Real-Time PCR Quantification of Chloroplast DNA Supports DNA Barcoding of Plant Species.

Species identification from extracted DNA is sometimes needed for botanical samples. DNA quantification is required for an accurate and effective examination. If a quantitative assay provides unreliable estimates, a higher quantity of DNA than the estimated amount may be used in additional analyses to avoid failure to analyze samples from which extracting DNA is difficult. Compared with conventional methods, real-time quantitative PCR (qPCR) requires a low amount of DNA and enables quantification of dilute DNA solutions accurately. The aim of this study was to develop a qPCR assay for quantification of chloroplast DNA from taxonomically diverse plant species. An absolute quantification method was developed using primers targeting the ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit (rbcL) gene using SYBR Green I-based qPCR. The calibration curve was generated using the PCR amplicon as the template. DNA extracts from representatives of 13 plant families common in Japan. This demonstrates that qPCR analysis is an effective method for quantification of DNA from plant samples. The results of qPCR assist in the decision-making will determine the success or failure of DNA analysis, indicating the possibility of optimization of the procedure for downstream reactions.

Forensic DNA Analysis of Wheat Flour as Commonly Used in White Powder Cases.

In the wake of terrorist attacks using anthrax and ricin, white powder is often encountered in cases of malicious mischief and terrorist threats. Wheat flour is a common white powder encountered in such criminal investigations. We used DNA analysis to investigate wheat flour samples for identification and discrimination as trace evidence. Species identification of commercially available wheat flour was carried out by sequencing a partial region of the ribulose bisphosphate carboxylase large subunit gene (rbcL). Samples were discriminated using short tandem repeat (STR) analysis. The rbcL sequences of all wheat flour samples were identical and showed a high level of similarity to known wheat (Triticum aestivum L.) sequences. Furthermore, flours had characteristic patterns in STR analyses, with specific cultivars showing distinctive patterns. These results suggested that the identification of wheat flour species is possible using rbcL sequencing, and that STR analysis is useful for discriminating between samples.

Effects of printing and ninhydrin treatment on forensic analysis of paper.

Paper is ubiquitous in human activities and can be found as evidence in the commission of many crimes such as threatening letters, deceptive advertisements and counterfeiting banknotes. To link the paper evidence to a source is a comparative process that is hampered when a blank paper is compared to a paper that has been submitted to printing or other treatments such as ninhydrin for the detection of fingermarks. During a forensic investigation, printed paper is analyzed with various instruments after fingerprint examination. In this study, the effects of printing and ninhydrin treatment on forensic paper examination of grammage, thickness, fillers, and pulp composition were studied. Grammage and thickness were increased by full-page double-sided printing, and grammage depended on the type of printer. The effects of printing on the analytical data about fillers and pulp composition were negligible, and ninhydrin treatment affected only paper thickness. These minor effects notwithstanding, the results indicate that conventional analytical methods used in forensic science for examining papers can be applied.

Potential utility of DNA sequence analysis of long-term-stored plant leaf fragments for forensic discrimination and identification.

This study examined the potential utility of DNA sequence analysis to discriminate and identify plant material in forensic investigations. DNA was extracted from plant leaf fragments of 11 species stored for 5 to 22 years after collection. The trnH-psbA intergenic spacer and 316 bp of the rbcL gene were successfully amplified and sequenced for all fragments except for the trnH-psbA spacer of one sample. All of the plant samples were discriminated in pairwise comparisons of the sequences. Using a combination of local and global genetic databases is likely to provide greater reliability in search results to identify forensic samples from sequence data.

Illegal route estimation of the seized illicit drug, methamphetamine, by the comparison of striation marks on plastic packaging films.

In Japan, the most common illicit drug is methamphetamine. It is possible to trace the origin of this drug by analyzing its organic and inorganic impurities and/or byproducts using several methods, such as GC, GC/MS, and inductively coupled plasma-mass spectrometry (ICP-MS). As reported here, one other method includes comparison of the striation lines of polymer sheet layers from packaging using a polarized light method. Other alternative methods include analyzing the heat sealer pattern, layer thickness surface characteristics, and/or components of polymer sheet layers using infrared spectroscopy. Several of these alternative methods were used to analyze the origins of 29 packages confiscated from three regions over a 1000 km distance in Japan. Results indicated that packages seized from different regions had some polymer sheet layers which contained striation lines and heat sealer patterns that were similar.

Guilty by his fibers: suspect confession versus textile fibers reconstructed simulation.

In one particular criminal case involving murder and theft, the arrested suspect admitted to the theft, but denied responsibility for the murder of the inhabitant of the crime scene. In his confession, the suspect stated that he found the victim's body when he broke into the crime scene to commit theft. For this report, the actual crime scene was reconstructed in accordance with the confession obtained during the interrogation of the suspect, and suspect behavior was simulated in accord to the suspect confession. The number of characteristic fibers retrieved from the simulated crime scene was compared with those of retrieved from the actual crime scene. By comparing the distribution and number of characteristic fibers collected in the simulation experiments and the actual investigation, the reliability of the suspect's confession was evaluated. The characteristic dark yellowish-green woolen fibers of the garment that the suspect wore when he entered the crime scene were selected as the target fiber in the reconstruction. The experimental simulations were conducted four times. The distributed target fibers were retrieved using the same type of adhesive tape and the same protocol by the same police officers who conducted the retrieval of the fibers at the actual crime scene. The fibers were identified both through morphological observation and by color comparisons of their ultaviolet-visible transmittance spectra measured with a microspectrophotometer. The fibers collected with the adhesive tape were counted for each area to compare with those collected in the actual crime scene investigation. The numbers of fibers found at each area of the body, mattress and blankets were compared between the simulated experiments and the actual investigation, and a significant difference was found. In particular, the numbers of fibers found near the victim's head were significantly different. As a result, the suspect's confession was not considered to be reliable, as a stronger contact with the victim was demonstrated by our simulations. During the control trial, traditional forensic traces like DNA or fingerprints were mute regarding the suspect's says. At the opposite, the fiber intelligence was highly significant to explain the suspect's behavior at the crime scene. The fiber results and simulations were presented at the court and the man was subsequently found guilty not only of theft and trespassing but also murder.

Forensic discrimination of match heads by elemental analysis with inductively coupled plasma-atomic emission spectrometry.

In arson and bombing cases, matches are often used as the ignition method. We have investigated the use of elemental analysis by inductively coupled plasma-atomic emission spectrometry to discriminate match heads used in arson cases. Six elements, magnesium, aluminum, calcium, iron, zinc, and barium, in match heads were detected after the match heads were dissolved in HNO3, and these elements were quantified in 8 wood stick matches and 5 paper stick matches by means of calibration curves prepared from standard sample solutions. Using this method, we were able to distinguish all the matches from one another both before and after combustion. The method has the potential to be very useful for resolving arson cases.

5-(4-Nitrophenyl)-2,4-pentadien-1-al (NPPD) used as a tracer by nonscientists at a simulated crime scene.

5-(4-Nitrophenyl)-2,4-pentadien-1-al (NPPD) was used as a tracer by nonscientists at a simulated crime scene. A card with both a plastic-coated smooth surface and a porous cellulose matrix paper surface was coated with a methanol solution containing 0.5mg/mL of NPPD. The card was touched with bare fingers and fingers covered by a cotton glove. A color-change protocol was then used to detect the presence of NPPD. The bare fingers or the fingers of gloves were swabbed with a cotton swab, or the parts of the glove that had touched the card were cut out. The swabs or the cloth pieces were dipped in methanol, a 0.1% methanol solution of naphthoresorcinol was added, and then concentrated hydrochloric acid was added. The observation of a red color at this point indicated a positive test. NPPD was easily observed in the experiments involving bare fingers, but no color change was observed from the swabbing of the cotton glove. However, when the cloth pieces cut from the fingers of the glove were subjected to the test, the red color was observed. In an attempt to enhance the sensitivity of the test, the volumes of the reagent solutions were reduced, but no improvement in sensitivity was obtained.

Discrimination of arsenous acids with comparison of trace elements contained by using synchrotron X-ray fluorescence spectrometry.

In the poisonous incident case occurred in Japan, clarification of the identity between seized poisons and retrieved from crime scene is strictly required from the court. In this case, arsenous acid was used as a poisonous material and, seized one from suspect's house was only twenty particles. The synchrotron X-ray fluorescence spectrometry by comparing the intensity ratios of L(alpha) line of four heavy metal, such as Bi, Sn, Sb, Se to K(beta) line of As was performed to overcome this problem. In this paper, the evaluation of this new method using 13 authentic arsenous acid samples, 4 of 13 were refined by Chinese method, 7 of 13 were refined by Japanese method (Sumitomo mining Co. Ltd. method), 2 of 13 were refined by German and Swiss method. As a result, by the comparison of the ratios of these four elements to As, these 13 samples were clearly classified to three products classes produced by different refining methods.

Evaluation of 5-(4-nitrophenyl)-2,4-pentadien-1-al (NPPD) as a tracer for shadowing pursuits.

The chemical compound 5-(4-nitrophenyl)-2,4-pentadien-1-al (NPPD), called "spy dust," was used in the Soviet Union as a shadowing pursuit, the act of following someone secretly, for investigating the activities of diplomatic personnel. It is also useful for counter-terrorism, and some criminal cases in the forensic science field. In this paper, it was synthesized and evaluated as a tracer for shadowing pursuits. The method for utilizing this reagent was very simple: it was dissolved in methanol (1 mg/mL) and sprayed on the restricted area. If the suspect was to enter this area or touched the sprayed material, NPPD was attached to the suspect's shoe surface or hands. The color examination was a two-steps process: first was the addition of 1 mL of a 0.1% naphthoresorcinol methanol solution to the methanol extracts of a methanol-contained cotton swab used to smear some surfaces of the suspect, and second, the addition of 1 mL of concentrated hydrochloric acid, which turned the solution dark red. The gamma max of the colored solution was 510 nm, measured by ultraviolet-vis spectroscopy. Detection limits for three methods were determined: a visual method (detection limit 100 ng/3 mL), an ultraviolet-visible spectrometric method (detection limit 10 ng/3 mL), and a selected-ion-monitoring gas chromatographic/mass spectrometric method (detection limit 300 pg/injection). The forensic utility of NPPD was demonstrated for two simulated cases: a theft case and a case where NPPD was used as a tracer to prove that an automobile had entered a restricted area. These examinations prove NPPD is a useful shadowing pursuit (spy dust) for the forensic science field.

Morphological and chemical analysis of magic mushrooms in Japan.

Morphological and toxicological analyses were performed on hallucinogenic mushrooms that are currently circulated in Japan. Scanning electron microscope (SEM) indicated a three-dimensional microstructures in the mushrooms. The complementary use of SEM with an optical microscope was effective for observing characteristic tissues, such as basidiomycetes, spores, cystidia and basidia. Hallucinogenic alkaloids were extracted with methanol and determined by high performance liquid chromatography (HPLC) with a UV detector set at 220 nm. The psilocin/psilocybin contents in Psilocybe cubensis were in the range of 0.14-0.42%/0.37-1.30% in the whole mushroom (0.17-0.78%/0.44-1.35% in the cap and 0.09-0.30%/0.05-1.27% in the stem), respectively. The hallucinogenic alkaloids in Copelandia were 0.43-0.76%/0.08-0.22% in the whole mushroom (0.64-0.74%/0.02-0.22% in the cap and 0.31-0.78%/0.01-0.39% in the stem). It thus appears that P. cubensis is psilocybin-rich, whereas Copelandia is psilocin-rich.