Forensic classification of ballpoint pen inks using high performance liquid chromatography and infrared spectroscopy with principal components analysis and linear discriminant analysis

Several varieties of blue ballpoint pen inks were analyzed by high performance liquid chromatography (HPLC) and infrared spectroscopy (IR). The chromatographic data extracted at four wavelengths (254, 279, 370 and 400 nm) was analyzed individually and at a combination of these wavelengths by the soft independent modeling of class analogies (SIMCA) technique using principal components analysis (PCA) to estimate the separation between the pen samples. Linear discriminant analysis (LDA) measured the probability with which an observation could be assigned to a pen class. The best resolution was obtained by HPLC using data from all four wavelengths together, differentiating 96.4% pen pairs successfully using PCA and 97.9% pen samples by LDA. PCA separated 60.7% of the pen pairs and LDA provided a correct classification of 62.5% of the pens analyzed by IR. The results of this study indicate that HPLC coupled with chemometrics provided a better discrimination of ballpoint pen inks compared to IR. The need to develop a suitable IR method for analysing blue ballpoint pen inks has been emphasized and it is hoped that the development of such a method would indeed provide a valuable tool for the non-destructive analysis of blue ballpoint pen ink samples for forensic purposes.     

A comparison of laser ablation inductively coupled plasma mass spectrometry,micro X-ray fluorescence spectroscopy,and laser induced breakdown spectroscopy for the discrimination of automotive glass

Laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS), micro X-ray fluorescence spectroscopy (μXRF), and laser induced breakdown spectroscopy (LIBS) are compared in terms of discrimination power for a glass sample set consisting of 41 fragments. Excellent discrimination results (> 99% discrimination) were obtained for each of the methods. In addition, all three analytical methods produced very similar discrimination results in terms of the number of pairs found to be indistinguishable. The small number of indistinguishable pairs that were identified all originated from the same vehicle. The results also show a strong correlation between the data generated from the use of µXRF and LA-ICP-MS, when comparing µXRF strontium intensities to LA-ICP-MS strontium concentrations. A 266 nm laser was utilized for all LIBS analyses, which provided excellent precision (< 10% RSD for all elements and < 10% RSD for all ratios, N = 5). The paper also presents a thorough data analysis review for forensic glass examinations by LIBS and suggests several element ratios that provide accurate discrimination results related to the LIBS system used for this study. Different combinations of 10 ratios were used for discrimination, all of which assisted with eliminating Type I errors (false exclusions) and reducing Type II errors (false inclusions). The results demonstrate that the LIBS experimental setup described, when combined with a comprehensive data analysis protocol, provides comparable discrimination when compared to LA-ICP-MS and μXRF for the application of forensic glass examinations. Given the many advantages that LIBS offers, most notably reduced complexity and reduced cost of the instrumentation, LIBS is a viable alternative to LA-ICP-MS and μXRF for use in the forensic laboratory.     

NACE Discrimination of Black Ballpoint Pen Inks

A non-aqueous capillary electrophoresis (NACE) method was developed for analyzing seven basic dyes contained in black ballpoint pen inks. Baseline separation of the studied compounds was achieved on a 57 cm × 75 μm capillary by using a non-aqueous solution composed of 1.0% acetic acid and 60 mM ammonium acetate in methanol medium. Based on the pattern of NACE electropherograms, 120 black ballpoint pens from different manufacturers were divided into six groups in terms of dye categories. Moreover, the black ballpoint pens from the same group may be further distinguished by cluster analysis based on the content of different dyes and some unknown peaks. Our results indicated that the developed NACE method is a credible warrant for investigation of the fraudulent documents.     

Forensic Identification of Dyes in Ballpoint Pen Inks Using LC–ESI–MS

The aim of this work was to develop a new technique using flow injection analysis combined with LC–ESI–MS which allows identification of dyes in ballpoint pen inks. A sample preparation procedure for the extraction of dyes from ballpoint pen strokes has been developed. The characteristic group of ions for each sample of 21 studied ballpoint pen inks corresponding to the present dyes has been determined using flow injection method. LC separation conditions for identified dyes have been optimized on reversed-phase sorbent based on silica gel. The best composition of the mobile phase for the dyes mixture LC separation was 0.1% aqueous formic acid and acetonitrile. Detection of dyes was carried out using mass spectrometry with electrospray ionization in positive and negative modes after reversed-phase liquid chromatography separation. Dye composition of ink was additionally confirmed comparing the data obtained from the literature. Flow injection analysis allows obtaining intensive ions of unknown strokes. It is difficult to get this information using only chromatographic methods, because dyes peak intensity can be low and noise of basic line high. Flow injection method allows distinguishing the analyzed 21 ballpoint pens by determining a characteristic set of dyes. The developed flow injection technique is very simple and quick. As a result, a novel approach for the identification of dyes in the ballpoint pen inks by flow injection analysis with LC–ESI–MS and UV detection without using standard dye samples has been established. It can be an effective alternative to the existing LC–DAD–MS and IR spectroscopy methods.     

Nondestructive classification and identification of ballpoint pen inks by Raman spectroscopy for forensic document examinations

As a non-destructive analytical method, Raman spectroscopy often provides insufficient information to identify or differentiate the ink used for the preparation of a questioned document. In this study, blue and black ballpoint pen inks deposited on paper substrate were examined in situ by conventional Raman spectroscopy. Inks were successfully classified based on the total number of prominent bands in Raman spectra. It was found that more than 90% of the samples of the same type and color could be differentiated visually using only Raman spectra, i.e. 94 and 95% for blue and black inks, respectively. As a result of this study, a flow chart has been constructed for blue and black ballpoint pen inks allowing their systematic identification. Raman spectroscopy proved to be a fast and precise technique for forensic ink analysis.     

Analysis of roller pen inks by capillary zone electrophoresis

The analysis of roller pen inks has become more and more important in fraudulent document examination because of the extensive use of roller pens in financial documents. Capillary electrophoresis with powerful resolution was applied for the analysis of roller pen inks. The experiment focused on the optimization of the separation of the extract from commercially available roller pen entries. A better separation electropherogram was obtained when a 20 mM borate buffer at pH 8.5 and a fused silica capillary with an inner diameter of 100 μm with a total length of 47 (40 cm to the detector window) were used. Five inks from roller pens of different manufacturers and countries were analyzed, and their electropherograms showed that most patterns are distinctly different from each other. Capillary with inner diameter of 100 μm increased the intensity of determination; therefore, color dyes were identified in the visible range and were able to provide more information for comparing types of roller pen inks.     

Characterization and forensic analysis of soil samples using laser-induced breakdown spectroscopy (LIBS)

A method for the quantitative elemental analysis of surface soil samples using laser-induced breakdown spectroscopy (LIBS) was developed and applied to the analysis of bulk soil samples for discrimination between specimens. The use of a 266 nm laser for LIBS analysis is reported for the first time in forensic soil analysis. Optimization of the LIBS method is discussed, and the results compared favorably to a laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) method previously developed. Precision for both methods was <10% for most elements. LIBS limits of detection were <33 ppm and bias <40% for most elements. In a proof of principle study, the LIBS method successfully discriminated samples from two different sites in Dade County, FL. Analysis of variance, Tukey’s post hoc test and Student’s t test resulted in 100% discrimination with no type I or type II errors. Principal components analysis (PCA) resulted in clear groupings of the two sites. A correct classification rate of 99.4% was obtained with linear discriminant analysis using leave-one-out validation. Similar results were obtained when the same samples were analyzed by LA-ICP-MS, showing that LIBS can provide similar information to LA-ICP-MS. In a forensic sampling/spatial heterogeneity study, the variation between sites, between sub-plots, between samples and within samples was examined on three similar Dade sites. The closer the sampling locations, the closer the grouping on a PCA plot and the higher the misclassification rate. These results underscore the importance of careful sampling for geographic site characterization.     

Analysis of Dye Components in Red Ballpoint Pen Inks by Nonaqueous Micellar Electrokinetic Chromatography with LIF Detection

An effective and economical nonaqueous micellar electrokinetic chromatography (NAMEKC) method was developed for the analysis of dye components with similar structure in red ballpoint pen inks. The new NAMEKC method using sodium dodecyl sulfate (SDS) micelle as pseudo-stationary phase in formamide/H₂O medium provided higher resolution and better selectivity than other conventional capillary electrophoresis methods. In this study, the experimental parameters including solvent medium, acid concentration, SDS concentration, separation voltage and temperature, were optimized. The optimized conditions were formamide/H₂O (90/10, v/v) with SDS at concentration of 100 mmol and 3.5 % acetic acid (v/v). The separation voltage and temperature were of 20 kV and 25 °C, respectively. Under the optimized conditions, a good separation of three red color dyes with similar structure was achieved. This method was successfully used to analyze several red ballpoint pen inks from different manufacturers with satisfactory result. Therefore, the proposed method could be useful for the discrimination of red ballpoint pen inks in questioned documents examination.     

Classification and dating of black gel pen ink by ion-pairing high-performance liquid chromatography

A novel approach for classification and dating of the black gel pen ink entries on document was developed based on ion-pairing high-performance liquid chromatography (IP-HPLC). Ninety-three black gel pens were collected and divided into two groups, dye-based and pigment-based, by preliminary solubility test. The chromatographic conditions for separation of the dye-based black gel pen inks were optimized and the dye components in inks were satisfactorily separated by using 40 mmol/L tetrabutylammonium bromide as ion-pairing reagent. According to the number and the chromatographic retention times of the main dye components, the 50 dye-based inks were categorized into four classes. The inks within a class can be further identified by the percentage of each dye component. The compositional changes of the dye components in the black gel pen ink entries on paper were investigated in light and natural aging conditions and it has been found that the dye components in the ink entries underwent obvious decomposition, and the decomposing extent of the dye components was related to the aging time. The results can provide scientific evidences for dating of the suspicious black gel pen ink entries on documents.     

Forensic comparative glass analysis by laser-induced breakdown spectroscopy

Glass samples of four types commonly encountered in forensic examinations have been analyzed by laser-induced breakdown spectroscopy (LIBS) for the purpose of discriminating between samples originating from different sources. Some of the glass sets were also examined by laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS). Refractive index (RI) measurements were also made on all glass samples and the refractive index data was combined with the LIBS and with the LA-ICP-MS data to enhance discrimination. The glass types examined included float glass taken from front and side automobile windows (examined on the non-float side), automobile headlamp glass, automobile side-mirror glass and brown beverage container glass. The largest overall discrimination was obtained by employing RI data in combination with LA-ICP-MS (98.8% discrimination of 666 pairwise comparisons at 95% confidence), while LIBS in combination with RI provided a somewhat lower discrimination (87.2% discrimination of 1122 pairwise comparisons at 95% confidence). Samples of side-mirror glass were less discriminated by LIBS due to a larger variance in emission intensities, while discrimination of side-mirror glass by LA-ICP-MS remained high.     

DATINK pilot study: An effective methodology for ballpoint pen ink dating in questioned documents

Establishing the approximate age of an ink entry from a questioned document is often a complicated task and a controversial issue in forensic sciences. Among the existing approaches, the analysis of solvents in ballpoint inks may be a useful parameter for determining the age of ink on paper. In recent years, several ink dating methods have been proposed. These methods have been based on the analysis of common ink solvents using gas chromatography/mass spectrometry (GC/MS) as the analytical platform. Despite these recent methods, several questions remain. The aim of this work was to develop an ink dating methodology (DATINK) for documents written by ballpoint pens based on the disappearance of volatile solvents from the ink entry. Multiple solid-phase microextraction (MHS-SPME) coupled to GC/MS was used to measure the solvents from ink entries made with four BIC^® ballpoint pens. The β parameter, the remaining fraction of the analyte in the system after one equilibration, corresponding to the successive extractions was considered for modelling a mathematical equation for later ink age dating. Preliminary tests of DATINK method showed that it was possible to detect the presence of ink solvents on documents up to the studied five years. The analyses of different real samples of known age were analyzed in terms of β values, which provided a mean relative error of 21%. The proposed use of β parameter for estimating the absolute age of ballpoint ink entries has shown promising results with a standard deviation of β ranging from 0.002 to 0.004.     

Multivariate classification of pigments and inks using combined Raman spectroscopy and LIBS

The authenticity of objects and artifacts is often the focus of forensic analytic chemistry. In document fraud cases, the most important objective is to determine the origin of a particular ink. Here, we introduce a new approach which utilizes the combination of two analytical methods, namely Raman spectroscopy and laser-induced breakdown spectroscopy (LIBS). The methods provide complementary information on both molecular and elemental composition of samples. The potential of this hyphenation of spectroscopic methods is demonstrated for ten blue and black ink samples on white paper. LIBS and Raman spectra from different inks were fused into a single data matrix, and the number of different groups of inks was determined through multivariate analysis, i.e., principal component analysis, soft independent modelling of class analogy, partial least-squares discriminant analysis, and support vector machine. In all cases, the results obtained with the combined LIBS and Raman spectra were found to be superior to those obtained with the individual Raman or LIBS data sets.     

显微共聚焦拉曼光谱结合群分析的黑色直液笔墨迹鉴别研究

黑色直液笔是一种新型书写工具,目前对该种笔墨迹的相关研究较少。为给文件检验工作中墨迹的分析提供新的参考依据,本实验使用显微共聚焦拉曼光谱技术,采集了30支不同品牌、型号的黑色直液笔墨迹光谱数据,进行Savitzky-Golay卷积平滑处理后,依据光谱图的拉曼位移及拉曼谱峰差异对墨迹进行初步分析。设置聚类方法为组间联接,区间距离测量方式为平方欧式距离,对采集的光谱数据进行群分析,将30支黑色直液笔墨迹样本分成了3类,并与品牌建立了相关联系;同时通过主成分分析验证了群分析的可靠性和准确性。研究表明,显微共聚焦拉曼光谱技术结合群分析方法可实现对黑色直液笔墨迹的无损分析及有效鉴别,该方法操作简便、结果准确,适用于法庭科学文件检验。     

Applicability of surface-enhanced resonance Raman scattering for the direct discrimination of ballpoint pen inks

In situ surface-enhanced resonance Raman spectroscopy (SERRS) with excitation at 685 nm is suitable for the direct discrimination of blue and black ballpoint pen inks on paper. For black inks, shorter excitation wavelengths can also be used. For blue inks, SERRS at 514.5 and 457.9 nm does not provide adequate discriminative power. At these excitation wavelengths, the SERRS signals of the Methyl Violet derivatives present in inks easily dominate the overall spectrum because of resonance enhancement and preferential interaction with silver sol particles. At 685 nm, this problem is not encountered as the Methyl Violet derivatives do not show resonance enhancement, while other components may still exhibit resonance. Thirteen blue and thirteen black ink lines were examined. For the blue and black inks, on the basis of the 685 nm SERR spectra, eight and six groups of spectra, respectively, could be distinguished. This discrimination largely agrees with information from thin layer chromatography (TLC) experiments, although some differences in group compositions are found. The in situ SERR spectra show good repeatability with regard to the Raman frequencies, band shapes and relative intensities of the spectral bands. However, absolute intensities cannot be used for discrimination purposes.     

Comparison of laser ablation-inductively coupled plasma-mass spectrometry and micro-X-ray fluorescence spectrometry for elemental imaging in Daphnia magna

Visualization of elemental distributions in thin sections of biological tissue is gaining importance in many disciplines of biological and medical research. Laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) and scanning micro-X-ray fluorescence spectrometry (micro-XRF) are two widely used microanalytical techniques for elemental mapping. This article compares the capabilities of the two techniques for imaging the distribution of selected elements in the model organism Daphnia magna in terms of detection power and spatial resolution. Sections with a thickness of 10 and 20 μm of the fresh water crustacean Daphnia magna were subjected to LA-ICP-MS and micro-XRF analysis. The elemental distributions obtained for Ca, P, S and Zn allow element-to-tissue correlation. LA-ICP-MS and micro-XRF offer similar limits of detection for the elements Ca and P and thus, allow a cross-validation of the imaging results. LA-ICP-MS was particularly sensitive for determining Zn (LOD 20 μg g⁻¹, 15 μm spot size) in Daphnia magna, while the detection power of micro-XRF was insufficient in this context. However, LA-ICP-MS was inadequate for the measurement of the S distributions, which could be better visualized with micro-XRF (LOD 160 μg g⁻¹, 5 s live time). Both techniques are thus complementary in providing an exhaustive chemical profiling of tissue samples.     

Mapping of lead,magnesium and copper accumulation in plant tissues by laser-induced breakdown spectroscopy and laser-ablation inductively coupled plasma mass spectrometry

Laser-Induced Breakdown Spectroscopy (LIBS) and Laser Ablation Inductively Coupled Plasma Mass Spectrometry (LA-ICP-MS) were utilized for mapping the accumulation of Pb, Mg and Cu with a resolution up to 200 μm in a up to cm × cm area of sunflower (Helianthus annuus L.) leaves. The results obtained by LIBS and LA-ICP-MS are compared with the outcomes from Atomic Absorption Spectrometry (AAS) and Thin-Layer Chromatography (TLC). It is shown that laser-ablation based analytical methods can substitute or supplement these techniques mainly in the cases when a fast multi-elemental mapping of a large sample area is needed.     

Laser induced breakdown spectroscopy on soils and rocks: Influence of the sample temperature,moisture and roughness

ExoMars, ESA's next mission to Mars, will include a combined Raman/LIBS instrument for the comprehensive in-situ mineralogical and elemental analyses of Martian rocks and soils. It is inferred that water exists in the upper Martian surface as ice layers, “crystal” water or adsorbed pore water. Thus, we studied Laser Induced Breakdown Spectroscopy (LIBS) on wet and dry rocks under Martian environmental conditions in the temperature range − 60 °C to + 20 °C and in two pressure regimes, above and below the water triple point. Above this point, the LIBS signals from the rock forming elements have local minima that are accompanied by hydrogen (water) emission maxima at certain temperatures that we associate with phase transitions of free or confined water/ice. At these sample temperatures, the plasma electron density and its temperature are slightly lowered. In contrast to powder samples, a general increase of the electron density upon cooling was observed on rock samples. By comparing the LIBS signal behavior from the same rock with different grades of polishing, and different rocks with the same surface treatment, it was possible to distinguish between the influence of surface roughness and the bulk material structure (pores and grains). Below the triple point of water, the LIBS signal from the major sample elements is almost independent of the sample temperature. However, at both considered pressures we observed a hydrogen emission peak close to − 50 °C, which is attributed to a phase transition of supercooled water trapped inside bulk pores.     

裂解气相色谱法分析蓝色中性笔字迹的书写墨水种类及墨迹随书写时间的变化规律

中性笔是当前比较流行的一种书写工具,其墨水多采用颜料作为色料成分,很难用水和有机溶剂进行提取,故无法利用分析圆珠笔油墨和水性笔墨水所采用的气相色谱或高效液相色谱等方法进行检验。目前在法庭科学领域内还没有一种有效的方法用于分析中性笔字迹的书写墨水。采用裂解气相色谱法分析了蓝色中性笔字迹的书写墨水,根据色谱峰的个数和保留时间将收集的65个样品分成3类,其中大部分样品属于具有铜酞菁颜料主要色谱峰的第二大类。色谱峰的保留时间和峰面积比值的分析结果表明所建立的分析方法重现性好且稳定。在此基础上,对蓝色中性笔墨迹随书写时间的变化规律进行了初步研究,得出了老化曲线。     

2-D analysis of Ge implanted SiO2 surfaces by laser-induced breakdown spectroscopy

2-D elemental distribution of Ge in silicon oxide substrates with differing implantation doses of between 3 × 10¹⁶ cm^− 2 and 1.5 × 10¹⁷ cm^− 2 has been investigated by Laser-Induced Breakdown Spectroscopy (LIBS). Spectral emission intensity has been optimized with respect to time, crater size, ablation depth and laser energy. Atomic Force Microscopy (AFM) and Scanning Electron Microscopy (SEM) coupled with Energy-Dispersive X-Ray Spectroscopy (EDX) have been utilized to obtain crater depth, morphology and elemental composition of the sample material, respectively. LIBS spectral data revealed the possibility of performing 2-D distribution analysis of Ge atoms in silicon oxide substrate. EDX analysis results confirmed that LIBS is capable to detect Ge atoms at concentrations lower than 0.2% (atomic). LIBS as a fast semi-quantitative analysis method with 50 µm lateral and 800 nm depth resolution has been evaluated. Results illustrate the potential use of LIBS for rapid, on-line assessment of the quality of advanced technology materials during the manufacturing process.     

New challenges and insights in the detection and spectral identification of organic explosives by laser induced breakdown spectroscopy

With the objective of detection and identification of explosives, different organic compounds, including aromatic nitrocompounds, RDX, anthracene, 2,4-diaminotoluene (DAT), 4-methyl-3-nitroaniline (MNA) and pentaerythritol (PENT) have been analyzed by laser induced breakdown spectroscopy (LIBS). To avoid the secondary ionization and to discriminate between the spectral contribution due to air from that of the compound in the plasma generated in air, the emission signatures from atomic lines (C at 247.9 nm, H at 656.3 nm, N at 746.8 nm and O at 777.2 nm) and molecular bands (CN at 388.3 nm and C₂ at 516.5 nm) have been investigated in plasmas generated in air and in helium. The different possible pathways leading to the observation of molecular emissions have been studied, together with a discussion of the most useful tools for the explosives discrimination. Moreover, the effect of the laser fluence on the atomic and molecular emissions and their relationship with the oxygen balance of an organic explosive is presented.