Methods for the analysis of hard dental tissues exposed to high temperatures

The preservation of teeth that have been exposed to a heat source is a key factor in order to keep their value as evidence. This research paper presents some methods under development for the analysis of dental evidence after exposure to high temperatures; it also establishes age related alterations of young and aged teeth which have been exposed to direct heat. A total of thirty permanent healthy teeth (young=15 and old=15) were submitted to direct heat (n=14) and to a controlled raise of temperature (n=16); they were embedded in polymethylmetacrilate. For the macroscopic study, blocks were cut in half, polished and their surfaces were digitalized. Subsequently, blocks were put in slides, worn away and observed through a photomicroscope. The study of the colorimetric pattern was done by Corel Photo Paint 12.0 and Image J. software. All teeth submitted to the gradual raise of temperature presented a degree of structural damage lower than those exposed to direct heat. The internal surface of young teeth exposed to direct heat showed lines of fracture oblique to the long axis of the tooth; in contrast, a reticular pattern was observed on aged teeth. All teeth exposed to direct heat showed some matte-black coloration areas, while those submitted to a gradual increment of temperature showed a greyish colour in the enamel and a chalky-white in the cementum. By using both thermal treatments in young teeth, the amelodentinal junction was lost, thus preserving the integrity of each tissue. In aged teeth, the separation of this junction was produced because of the breakdown of the dentine. Microscopically, in teeth exposed to direct heat, it was neither possible to differentiate the dentin from the cementum, nor to distinguish the incremental lines of the latter; while in those submitted to a controlled increment of temperature, the corresponding morphology in the tissues was observed. A significant statistical difference was obtained for black colour among young and aged teeth exposed to direct heat, as well as among the means of all colours obtained from the teeth exposed to both treatments. The structural changes observed allow locating a tooth in the studied age groups. Our results show the viability of the applied method for the processing of samples, allowing their preservation as evidence during an indefinite time.     

The application of amino acid racemization in the acid soluble fraction of enamel to the estimation of the age of human teeth

Estimation of age-at-death for skeletonised forensic remains is one of the most significant problems in forensic anthropology. The majority of existing morphological and histological techniques are highly inaccurate, and show a bias towards underestimating the age of older individuals. One technique which has been successful in forensic age estimation is amino acid racemization in dentine. However, this method cannot be used on remains where the post-mortem interval is greater than 20 years. An alternative approach is to measure amino acid racemization in dental enamel, which is believed to be more resistant to change post-mortem. The extent of amino acid racemization in the acid soluble fraction of the enamel proteins was determined for modern known age teeth. A strong correlation was observed between the age of the tooth and the extent of racemization. No systematic bias in the direction of age estimation errors was detected. For the majority of teeth analyzed, the presence of dental caries did not affect the results obtained. In a minority of cases, carious teeth showed a higher level of racemization than would be expected given the age of the individual. These results indicate that amino acid racemization in enamel has the potential to be used in age estimation of skeletal remains.     

Age estimation methods using anthropological parameters on human teeth-(A0736)

The research was conducted on the 160 intact extracted human teeth with one and two roots of the known age and sex. The teeth were disinfected, dried and X-rayed. After that the section of the longitudinal cut through the teeth was performed in order to facilitate monitoring of all tissues and morphological characteristics of the teeth. The age was determined in three ways: Method 1 [G.Bang, E. Ramm, Determination of age in humans from root dentin transparency, Acta Odontol. Scand. 28 (1970) 3-35]--analysis of the translucency of the root dentine, Method 2 [S. Kvaal, T. Solheim, A non-destructive dental method for age estimation, J. Forensic Odonto-stomatol. 12 (1994) 6-11]--analysis of the root and the root canal from the X-ray, Method 3 [G. Johanson, Age determination from human teeth, Odontol. Revy. 22 (1971) 1-126]--analysis of six parameters on each teeth. All data were subject to the correlation and regression analysis which showed the following: all of the three applied methods were in the significant correlation with the real age, and the best of them proved to be Method 3 where the coefficient of correlation was 0.85, p<0.001. The teeth of the maxilla are more convenient for the age determination than the teeth of mandible. They are in the significant strong correlation with the known real age, and in Method 3, the coefficient of correlation is 0.78, p<0.001. Age determination of the teeth with two roots is in significant correlation with the known real age p<0.001 in relation to the determined age on the teeth with one root. The results show that sex too, is in significant correlation with the real age, p<0.001. In practice, the methods used and the results achieved in this research have been enabling the dental age estimation of human remains from mass graves after the 1991 war in Croatia.     

Objective measurement of dental color for age estimation by spectroradiometry

Age-related changes in tooth color have been described previously, however, the use of dental color for age estimation in forensic odontology has been limited due to the difficulty of measuring color objectively. This study presents an objective method for determining dental color to estimate the age of an individual. Dentine color in 250 teeth from patients ranging in age from 10 to 89 years was determined by spectroradiometry. Color measurements were performed as suggested in the CIE 1931 (International Commission on Illumination). Chromaticity coordinates (x, y, z), luminance (Y), whiteness index (WIC, Z%, WIC) and yellowness index (YI) were obtained. Correlations between these colorimetric variables and aging were established by linear regression analyses. All the variables fit the mathematical model with correlation coefficients ranging from 0.53 to 0.75. This method of color measurement produced an expected associated error of calibration averaging 13.7 years about the mean estimated values, at a 70% level of confidence. Two different multiple regression models for dental age estimation were tested, and variables that made the greatest contributions to age calculation were identified. To determine the effect of postmortem interval on tooth color and its influence in age estimation, 37 teeth obtained from human skeletal remains buried during an interval ranging from 21 to 37 years were also studied. In this material, the correlation between age and dental color measured by spectroradiometry was weaker than in fresh extracted teeth.It is concluded that determination of dentine color by spectroradiometry is a potentially useful objective method to estimate age in forensic studies in combination with other methods.     

X‐ray Scattering Evaluation of Ultrastructural Changes in Human Dental Tissues with Thermal Treatment

Micro‐ and ultrastructural analysis of burned skeletal remains is crucial for obtaining a reliable estimation of cremation temperature. Earlier studies mainly focused on heat‐induced changes in bone tissue, while this study extends this research to human dental tissues using a novel quantitative analytical approach. Twelve tooth sections were burned at 400–900°C (30‐min exposure, increments of 100°C). Subsequent combined small‐ and wide‐angle X‐ray scattering (SAXS/WAXS) experiments were performed at the Diamond Light Source synchrotron facility, where 28 scattering patterns were collected within each tooth section. In comparison with the control sample, an increase in mean crystal thickness was found in burned dentine (2.8‐fold) and enamel (1.4‐fold), however at a smaller rate than reported earlier for bone tissue (5–10.7‐fold). The results provide a structural reference for traditional X‐ray scattering methods and emphasize the need to investigate bone and dental tissues separately to obtain a reliable estimation of cremation temperature.     

Heat Induced Changes to Dental Resin Composites: A Reference in Forensic Investigations?*

The objective was to investigate color change and surface damage in dental resin composites exposed to high temperatures over different time intervals for comparative purposes. Samples were prepared using two resins - Z100(R) (R1) and Charisma (R2), heated at the following temperatures: 200 degrees C, 400 degrees C, 600 degrees C, 1000 degrees C, for 15, 30 and 45 min (n = 104 for each resin sample). Color (DeltaE) and brightness (DeltaL) changes were analyzed by spectrophotometry using the CIE Lab system and surface changes by Scanning Electron Microscopy (SEM). R1 showed more intense color changes after heat exposure than R2. DeltaL values were found to be the best parameter for evaluation of light and color change. A biphasic pattern after thermal exposure was detected, from dark brown to light white. SEM showed more intense alterations in R2 than in R1. These results indicate that the parameters observed in both resins are useful as a guide in forensic analyses.     

Model of age estimation based on dental factors of unknown cadavers among Iranians

Two hundred and ten cadavers ranging in age from 25 to 60 were studied. Thirty subjects from each five-year interval were selected, and data were collected using the mandibular anterior and premolar teeth after extraction. Sections ranging from 0.5 to 1.0 mm were measured by a stereo microscope with a precision of 0.1 mm. The following factors were determined: attrition, periodontosis, root resorption, secondary dentine apposition, cementum apposition, and translucency of the root. Statistical analysis used the sum of ranks of the dental factors as an independent variable in a linear regression model to estimate the age of the cadaver. Among the different mandibular teeth, the sum of ranks of the first premolar factors had the best correlation coefficient with age. The sum of the dental factors presented a better model than each of the factors alone. The first premolar is recommended as the first step in the estimation of age.     

The use of quantitative light-induced fluorescence (QLF) to identify composite restorations in forensic examinations

There has been a large increase in the number of tooth colored restorations "white fillings" placed in recent years. An increased demand from the public for more aesthetic dental restorations causes a potential problem for forensic dentists who may find the fillings difficult to identify and hence include in postmortem odontograms. This has implications for the accuracy of dental identifications, particularly in situations where limited time is available for postmortem identification, e.g., mass casualty incidents. A new method for the detection of composite restorations is presented. Quantitative Light-induced Fluorescence (QLF) is a technique currently employed to detect small changes in enamel mineral content. An experiment was conducted to determine if the technique would afford a greater degree of contrast between composite and enamel and thus enable the accuracy of composite identification in enamel. Twenty-four previously extracted human premolars were gently cleaned with pumice and wet-and-dry paper. Twelve were subsequently randomly selected and restored on their buccal surfaces with Spectrum (a composite) following manufacturer's instructions. No attempt was made to color match the teeth and all were filled with shade B3. Twelve teeth were left unrestored. QLF and normal white light images were taken of both restored and non-restored surfaces with teeth wet and then dried. Ten forensic dentists were asked on two separate occasions (one month between each attempt) to indicate whether or not they thought the surface was: a) restored or b) unrestored. Results indicate that forensic dentists detected a significantly higher proportion (p<0.005) of filled surfaces with QLF.     

Differences in morphological age-related dental changes depending on postmortem interval.

Morphological methods for age estimation from teeth have been developed and applied to samples without taking the postmortem interval into consideration. We studied differences in morphological age-related changes between fresh extracted teeth and teeth from human skeletal remains in order to develop appropriate dental age estimation methods according to the time after death. Forty-three permanent teeth from dental patients were compared to 37 teeth obtained from human skeletal remains with a postmortem interval from 21 to 37 years. Morphological age-related changes were investigated by measuring variables on intact and half-sectioned teeth. A new computer assisted image analysis procedure to avoid subjectivity was developed to measure variables in sectioned specimens. Dental color, translucency length, attrition, cementum apposition, and secondary dentin showed higher values in teeth from human skeletal remains than in fresh extracted teeth. Variables obtained by morphometric analysis of computer-generated images (tooth length, tooth width, root length, and root area) showed higher values in fresh extracted teeth than in teeth from skeletal remains. The postmortem interval affects age-related morphological changes, and therefore different methods should be used for teeth of unknown postmortem interval.     

Postmortem red coloration of teeth. A retrospective investigation of 26 cases

Postmortem red coloration of teeth (pink teeth) was found in 26 of 119 consecutive autopsy or forensic dental identification cases during 1973-1984. The present study confirms that red coloration of teeth may occur some time after death under conditions that promote putrefaction and adipocere formation. The red coloration is due to hemolysis after exudation of hemoglobin derivatives through the dentine tubules in cases in which blood has accumulated in the head, either due to congestion or by a postmortem head-down position, as after submersion in water.     

Radiocarbon analysis of dental enamel and bone to evaluate date of birth and death: perspective from the southern hemisphere

Radiocarbon analysis was conducted on dental enamel, cortical bone and trabecular bone from four human adults from Andean Peru with known birth dates and death dates, the latter all falling within the modern bomb-curve period. Radiocarbon results were compared with known tropospheric values from the southern hemisphere. The results demonstrate that dental enamel can provide useful information regarding birth dates in consideration of known formation times of the specific teeth examined if the radiocarbon values fall within the modern bomb-curve period. Trabecular bone radiocarbon values are closer to the tropospheric values at the date of death than are values from cortical bone reflecting relative differences in the rate of bone remodeling in those tissues. Age at death also represents an important factor.     

Color Stability of Dental Restorative Materials Submitted to Heat Sources,for Forensic Purposes

During postmortem examination of the dental arches of carbonized victims, dental restorative materials may be found. The aim of this study was to evaluate the effect of heat source action on the color stability of composite resin (CR) and glass ionomer cement (GIC) restorations, to discriminate between them and compare with antemortem dental data. Sixty bovine teeth (30 CR and 30 GIC) were prepared (6 × 6 × 2 mm) and separated into groups (n = 10). The color readouts were taken by spectrophotometer, before and after heat action (100°C, 200°C, 300°C), in an oven for 15 min. There were color alterations for all coordinates (ΔE, ΔL*, Δa* eΔb*) for both materials. GIC presented greater change. The authors concluded that it is possible to distinguish between the materials by the color changes analyzed by instrumental method, helping victim identification.     

Changes in perikymata and their significance to a postmortem dental identification

Perikymata are wavelike features that are normally found in the surface enamel of teeth. These structures are known to undergo regressive changes due to gradual attrition, mechanical abrasion, or chemical erosion. Forensic science has suggested that significance be placed on some forms of their loss as indicators of age, oral habits, and alignment of individual teeth. The dental identification of victims of a Vietnam War military aircraft crash is presented. The unusual pattern of perikymata change on two of the teeth suggests the antemortem existence of a removable partial denture. These findings support the exclusionary conclusions.     

A modified Gustafson method of age estimation from teeth

Modifications to the Gustafson method of age estimation from teeth are described. They are designed to minimise the subjective nature of the scoring technique and to quantify the regressive changes in the enamel, dentine and cementum. Randomly selected teeth from 25 Indians were examined by the Gustafson method and by our own modification. It was shown that our modified method resulted in more accurate and reproducible age estimations than did the original method.     

Multiple regression models for age estimation by assessment of morphologic dental changes according to teeth source

The aims of this study were twofold: (1). to measure parameters that contribute significantly to estimates of dental age, using a combination of classic methods and a computer-assisted image analysis procedure to avoid the bias inherent in observer subjectivity; and (2). to develop new mathematical regression models for age prediction according to postmortem interval. Two different populations were studied. Forty-three permanent teeth (Group I), extracted for valid clinical reasons, were taken from patients 25-79 years of age. The other population group (Group II) was composed of 37 healthy erupted permanent teeth obtained from human skeletal remains (age 22-82 years) with a postmortem interval ranging from 21 to 37 years. Morphologic age-related changes were investigated by measuring variables on intact and half-sectioned teeth. Multiple regression analyses were performed with age as the dependent variable for each sample source. In fresh extracted teeth, the variables that made the greatest contributions to predictions of age were dental attrition, dentin color, and translucency width, the latter measured with a computer-assisted image analysis method. In teeth from human skeletal remains, the variables that made the greatest contributions to age calculation were cementum apposition, pulp length measured by computer-assisted image analysis, dental attrition, root translucency, and dental color. We conclude by recommending different regression models to calculate age depending on the postmortem interval.     

A Calibration Test for Latent Fingerprint Development on Thermal Paper

A calibration test is described for monitoring the operation of equipment used to develop latent fingerprints on thermal paper by the application of either controlled or uncontrolled heat. A working solution of a water/glycerol emulsion and butylene glycol is applied to thermal paper by means of either a vinyl stamp and pad, or a marker pen. Varying the amount of butylene glycol enables the thermal paper to change color at different temperatures between approximately 40 and 60°C, which is below the normal color change temperature of the paper. The described test may be used to verify the correct operation, at different temperatures, of a controlled heat source during and after fingerprint development (such as the Hot Print System) or to monitor the paper temperature with an uncontrolled heat source (such as a warm air blower), thereby avoiding unintentional coloring of the entire paper surface.     

alpha-L-fucosidase phenotyping in human tissues, dental pulps and hair roots

Phenotypes of alpha-L-fucosidase (Fu) were demonstrated from tissue samples of spleen, liver, lung and kidney stored for a few weeks at room temperature. Activity was fairly low in pancreas, heart, muscle, skin and brain, and typing was not reliable after 1 week storage. Fu types were detectable from dental pulp tissue of up to 1 week storage. Activity was present in hair root cells, but was extremely low. The results show that the Fu typing may be applicable to individual identification of organ tissues and teeth.     

Comparative Analysis of the Effects of Heat on the PCR‐Amplification of Various Sized DNA Fragments Extracted from Sus Scrofa Molars*

Abstract: This study examined the effects of heat on the amplification of DNA from the dental pulp of Sus scrofa molars and investigated the protection afforded to the pulp tissue by the dental enamel, alveolar process, and soft tissue of the head. Segments of defleshed maxilla and mandible encasing the first molar (n = 60) were subject to a range of temperatures for 15 min. Dental pulps were retrieved. Amplifications using three‐primer and four‐primer multiplexes showed no degradation of the largest fragment following exposure to 450°C. Amplifications in the three‐primer multiplex (283 bp) were successful following exposure to 525°C in maxillary samples only. This study revealed the enamel density of maxillary molars to be greater than mandibular molars in Sus scrofa. Following incineration of intact heads for 15 min (n = 10) and 1 h (n = 4) at an average temperature of 625°C, amplifications of the largest fragment (450 bp) were successful from both maxillary and mandibular teeth.     

Dental CT imaging as a screening tool for dental profiling: advantages and limitations

The use of dental processing software for computed tomography (CT) data (Dentascan) is described on postmortem (pm) CT data for the purpose of pm identification. The software allows reconstructing reformatted images comparable to conventional panoramic dental radiographs by defining a curved reconstruction line along the teeth on oblique images. Three corpses that have been scanned within the virtopsy project were used to test the software for the purpose of dental identification. In every case, dental panoramic images could be reconstructed and compared to antemortem radiographs. The images showed the basic component of teeth (enamel, dentin, and pulp), the anatomic structure of the alveolar bone, missing or unerupted teeth as well as restorations of the teeth that could be used for identification. When streak artifacts due to metal-containing dental work reduced image quality, it was still necessary to perform pm conventional radiographs for comparison of the detailed shape of the restoration. Dental identification or a dental profiling seems to become possible in a noninvasive manner using the Dentascan software.     

PGD phenotyping in bloodstains, organ tissues, dental pulps and hair roots by isoelectric focusing

Polymorphism of PGD was investigated in bloodstains, organ tissues, dental pulps, hair roots and semen by isoelectric focusing. This technique provided much higher resolution of PGD isoenzymes than starch gel electrophoresis. Phenotyping was possible from bloodstains for 5 weeks, from organ tissues (except pancreas) for 1-3 weeks, from dental pulps for 2 weeks and from hair roots for 2 weeks when they were stored at room temperature. The method is simple, rapid, reliable and therefore useful in medicolegal individualization of bloodstains, organ tissues, teeth and hairs.