A RESEARCH PROJECT TO ACQUIRE THE ACADEMIC TITLE OF
MASTER
OF
SCIENCE
IN
FORENSIC SCIENCE
FROM
UNIVERSITY OF AMSTERDAM
Done by:
S. DIVYA
Student ID:
11390476
Supervisor:
Drs. Ing. TRISTAN KRAP
Examiner:
Prof. dr. R.J. (ROELOF-JAN) OOSTRA
Research Institute:
Amsterdam Universitair Medische Centra (UMC)
Number of Credits:
36
Project Duration:
05/03/2018 – 27/09/2018
Date of submission:
20/09/2018
HIT OR BURN? –
DIFFERENTIATING BLUNT FORCE TRAUMA FRACTURES FROM
HEAT-INDUCED FRACTURES IN BONES
1 TABLE OF CONTENTS
1. ABSTRACT ... 3
2. INTRODUCTION ... 4
3. MATERIALS AND METHODS ... 8
3.1. Sample material ... 8
3.1.1. Sample preparation ... 8
3.1.2. Confounding factors ... 8
3.2. Pilot Study... 8
3.2.1. Pilot study (i): BFT generation ... 8
3.2.2. Pilot study (ii): Burning temperature and duration ... 9
3.3. Main study ... 10
3.3.1. BFT production ... 11
3.3.2. Maceration ... 11
3.3.3. Burning... 11
3.4. Observations: Macroscopic and microscopic analyses ... 12
3.4.1. Fracture visualization ... 13 3.4.2. Measurement of angles ... 13 3.4.3. Colourimetric analysis ... 13 3.5. Statistical analysis ... 14 4. RESULTS ... 16 4.1. Pilot study ... 16 4.1.1. (i): BFT generation ... 16
4.1.2. (ii): Burning temperature and duration ... 17
4.2. Main study ... 18
4.2.1. Post-BFT features (Pre-Maceration and Pre-Burning) ... 18
4.2.2. Post-Maceration (Group B) ... 19
4.2.3. Post-Burning (Group A and C)... 20
4.2.4. Comparison of BFT and HIF features between groups ... 23
4.2.5. Colourimetric analysis ... 24
4.2.6. Dimensional changes ... 26
5. DISCUSSION ... 28
5.1. Experimental design ... 28
5.2. Post-BFT fracture features ... 30
5.3. Features from unmacerated bone vs features from macerated bones ... 31
5.4. Post-burning features ... 31
5.5. Colourimetry findings ... 32
5.6. Dimensional changes ... 33
5.7. Limitations ... 34
5.8. Recommendations and scope for future research ... 34
6. CONCLUSION ... 35
7. ACKNOWLEDGEMENT ... 36
8. BIBILIOGRAPHY ... 37
2 LIST OF FIGURES
FIGURE 1:THE STRESS-STRAIN CURVE.IN BONES, FIRSTLY, ELASTIC DEFORMATION HAPPENS WHEN BONE RESUMES ITS ORIGINAL SHAPE AFTER
STRESS IS GONE. ... 5
FIGURE 2:DIFFERENT TYPES OF HIFS- A) LONGITUDINAL, B) CURVED TRANSVERSE, C) STRAIGHT TRANSVERSE, D) PATINA, E) DELAMINATION. ... 6
FIGURE 3:CUSTOM-MADE PENDULUM-LIKE CONTRAPTION TO PRODUCE FRACTURES. ... 9
FIGURE 4:PLACEMENT OF BONES ON THE CUSTOM-MADE PENDULUM-LIKE CONTRAPTION TO PRODUCE FRACTURES. ... 9
FIGURE 5:BOX PLOT SHOWING THE DISTRIBUTION OF BONE SAMPLES IN EACH STUDY GROUP, BASED ON AGE (LEFT), WEIGHT (CENTER) AND LENGTH (RIGHT) OF RESPECTIVE CADAVERS. ... 10
FIGURE 6:MACERATION SET-UP.. ... 11
FIGURE 7:SCHEMATIC REPRESENTATION OF THE ROOM IN FIREHOUSE THAT WAS USED FOR BURNING THE BONES. ... 11
FIGURE 8:BURNING EXPERIMENT SET-UP. ... 12
FIGURE 9:ANGLE MEASUREMENT USING IMAGEJ. ... 13
FIGURE 10:SECTIONS USED TO MEASURE L-B VALUES ... 14
FIGURE 11:EXAMPLE OF MICROSCOPIC OBSERVATION OF SHARPNESS PRE-MACERATION AND POST-MACERATION FOR GROUP B BONES... ... 20
FIGURE 12:DEPICTS THE PRESENCE OF BOTH ROUGHNESS AND SOME SMOOTHNESS (AT SLOPED/CURVED EDGES) IN BURNED BONE FROM GROUP C.. ... 21
FIGURE 13:DENOTES THE DIFFERENCE IN MARGINS APPEARANCE AND COLOURATION OF EDGES FOR SITUATIONAL FRACTURE AND BFT-FRACTURE. . ... 22
FIGURE 14:DEPICTS THE SAME PLANE OF MARGINS FOR HIFS AND ELEVATION OF MARGIN ON ONE SIDE OF SITUATIONAL FRACTURE.. ... 22
FIGURE 15:PHOTOGRAPHS OF GROUP A(LEFT) AND GROUP C(RIGHT) BONES, TAKEN AFTER BURNING, USING MIRROR-REFLEX CAMERA. ... 24
FIGURE 17:GRAPH SHOWS TEMPERATURE RANGES ACROSS EACH PORTION OF BONE FOR GROUP A, BASED ON THE UNPUBLISHED COLOURIMETRIC MODEL.. ... 25
FIGURE 16:GRAPH SHOWS TEMPERATURE RANGES ACROSS EACH PORTION OF BONE FOR GROUP C, BASED ON THE UNPUBLISHED COLOURIMETRIC MODEL.. ... 25
FIGURE 18:BOXPLOT FOR SHRINKAGE OF BONES GROUP C(POST-BURNING) FOR EACH DIMENSIONAL PARAMETER. ... 26
FIGURE 19:BOXPLOT FOR EXPANSION OF BONES GROUP C(POST-BURNING) FOR EACH DIMENSIONAL PARAMETER. ... 27
LIST OF TABLES TABLE 1:SOME MAJOR FEATURES OF TRAUMATIC AND HEAT-INDUCED FRACTURES AS WELL AS COEXISTENCE OF BOTH, AS OBSERVED IN FORMER STUDIES.. ... 4
TABLE 2:THE VARIOUS FEATURES EMPLOYED IN CURRENT STUDY AND THE CORRESPONDING FORMER STUDIES, WHICH INVESTIGATED FRACTURES USING THESE FEATURES. ... 7
TABLE 3:GROUPING OF FOREARM BONES, EACH GROUP WITH EQUAL NUMBER OF ULNA AND RADIUS.. ... 10
TABLE 4:INDICATES THE DISTRIBUTION OF BONES FROM MALES AND FEMALES (RIGHT) AND RADIUS/ULNA (LEFT) IN EVERY GROUP. ... 10
TABLE 5:INTERGROUP COMPARISONS AND THE DIFFERENT OUTCOMES IN RELATION TO SPECIFYING FEATURES OF EACH FRACTURE TYPE. ... 12
TABLE 6:DEPENDENT AND INDEPENDENT VARIABLES ASSOCIATED WITH THE STUDY ... 14
TABLE 7:SCORING LABELS FOR EACH FRACTURE FEATURE.. ... 15
TABLE 8:OBSERVATIONS FROM THE PILOT STUDY TO IDENTIFY THE IDEAL ANGLE FOR THE GENERATION OF BFT FRACTURE.. ... 16
TABLE 9:OBSERVATIONS FROM THE PILOT STUDY TO IDENTIFY THE IDEAL TEMPERATURE AND DURATION OF BURNING FOR THE INVERSION TO CALCINATION STAGE TO OCCUR. ... 18
TABLE 10:ANOVA AND KRUSKAL WALLIS TEST RESULTS FOR POST-BFT–GROUP B VS GROUP C. ... 19
TABLE 11:ANOVA AND KRUSKAL WALLIS TEST RESULTS FOR GROUP B– UNMACERATED VS MACERATED. ... 20
TABLE 12:ANOVA AND KRUSKAL WALLIS TEST RESULTS FOR POST-BURNING –GROUP A VS GROUP C. ... 23
TABLE 13:ANOVA AND KRUSKAL WALLIS TEST RESULTS FOR GROUP C– PRE-BURNING VS POST-BURNING. ... 23
TABLE 14:ANOVA AND KRUSKAL WALLIS TEST RESULTS FOR POST-BURNING GROUP A VS BFT ONLY GROUP B. ... 23
TABLE 15:ANOVA AND KRUSKAL WALLIS TEST RESULTS FOR BFT ONLY GROUP B VS POST-BURNING GROUP C. ... 24
TABLE 16:KRUSKAL WALLIS TEST RESULTS FOR THE TEMPERATURE DISTRIBUTION (CALCULATED USING COLOURIMETRY) OF GROUP A VS GROUP C (POST-BURNING). ... 26
3
1. ABSTRACT
Trauma interpretation from burned remains, being a persisting challenge, affects events reconstruction in forensic casework. The similarity, the co-existence and overlapping of features of traumatic fractures and heat-induced fractures (HIFs) accounts for this problem. Scarce literature exists for this issue, particularly for blunt force trauma (BFT) and these studies utilized animal bones. To address this problem, this project aims to distinguish the characteristics of BFT-fractures (focused only on complete fractures) and HIFs (and situational fractures) in burned bones. Defleshed fresh-frozen human cadaveric forearm bones were subjected to BFT (using pendulum-like contraption) and/or burning (wooden pyre in firehouse). The resulting fractures were scrutinized using a checklist of features (fracture morphology, colour and dimensional changes) derived from prior studies to determine these differences. Macro- and microscopic analyses were done. Colour changes were assessed using a colourimetric model. Traumatic fractures showed rough fractures edges with some smooth curved/slanted regions while HIFs displayed smooth surfaces, post-burning. Some longitudinal HIFs and longitudinal situational fractures mimicked the fracture lines of traumatic fractures, which could lead to misidentification of fracture type. Other overlapping features between the BFT-fractures and HIFs (and situational fractures) include smoothness seen in both fracture edges, situational fracture outlines being similar to transverse BFT-fracture outlines and fracture angles below 90. Interestingly, situational fracture margins were elevated on one side whereas HIFs penetrated into the medullary cortex. The former also showed even richly-coloured edges while BFT-fractures had uneven discolouration of fracture edges. No distinct trend was observed between fracture presence/formation, colouration and temperature range. Most of the observed features agreed with former studies. This study shows the effectiveness of a checklist of both quantitative and qualitative features for fracture differentiation and successful analysis using a stereomicroscope, a simpler and cheaper technique. Since fracture formation and/or alteration during burning are influenced by inter-individual variability and many environmental factors, further research is needed to understand the above differences. Conclusively, differentiation of traumatic fractures from HIFs is possible through meticulous analysis. However, experimental results may not always be applicable to forensic casework as they are not entirely representative of forensic case conditions.
4
2. INTRODUCTION
Fire-related incidences are common globally and nearly all fires have a forensic context – accidental (for example vehicular accidents and house fires from short-circuits) or intentional causes (like homicides) [1]. The latter constitute concealment of offences and the ensuing evidence including human bodies. In such cases, it is critical for investigators to determine presence of any perimortem trauma from the remains and allow accurate scenario formulations and events reconstruction and thus, relevant offences with lawfully correct sentences can be determined in court [2]. However, a major challenge in trauma interpretation from thermally-affected bodies lies in differentiation of mechanical (traumatic) fractures from heat-induced fractures (HIFs) [3,4]. This difficulty arises from characteristics of HIFs mimicking or being very similar to those of traumatic-fractures [1,3,5]. HIFs can also co-exist with BFT-fractures – either as independent fracture paths or merge with each other [6] (see Table 1 for these features and studies that investigated them). Since both types of fracture are multifactorial processes, it is hard to replicate exact casework conditions in the laboratory. Thus, extensive empirical research for this topic is needed [7]. Moreover, there is no standardization of clear descriptions of fracture-features, affecting the precise applicability of previous empirical data to casework [8]. The fracture distinction requires the understanding of (i) biomechanics of bone fractures and effect of fire on bones and (ii) coexistence of traumatic and heat-induced fractures (HIFs) on bones [9]. The aim of this project is to effectively (i) consider, (ii) to investigate and (iii) determine characteristics that differentiate mechanical fractures from HIFs, whilst addressing the aforementioned difficulties.
FORMER STUDIES
TRAUMATIC FRACTURES
HEAT-INDUCED FRACTURES COEXISTENCE OF BOTH TRAUMATIC AND HEAT-INDUCED FRACTURES Herrmann & Bennett, 1999 Greater definition of bony microstructure Larger fragments of bone Mostly smooth fracture surfaces
Generally smooth fracture surface Smaller fragments of bone due to
heat-caused fragmentation Transverse fractures with 90
fracture angles are common Longitudinal/oblique fracture
surface is typically rough with sectioned haversian/vascular canals
Longitudinal fracture propagation is similar and common to both trauma and burning
Traumatic fractures become rough and irregular with cleanly sectioned haversian/vascular canals Macoveciuc, Márquez-Grant, Horsfall & Zioupos, 2017 Sharp or jagged fracture outline with regular and smooth surface
Significant depth of fracture, whereby majority penetrates the entire cortex
Well-defined and sharp outline, smooth surface
Shrinkage of bone present
Sharpness of traumatic fracture outline lost with fracture surface becoming rough and irregular
Longitudinal and transverse fracture from both trauma and heat-induced show similar morphology
No alteration of traumatic fracture general pattern Curved transverse fractures are
specific to heat-induced fractures
Bone shrinkage present Pope & Smith,
2004 Impact caused by weapon seen in fractures as sharp margins, bone depressions and inward crushing
Increased bone brittleness
Colour changes observable Traumatic margins deformed, bevelled, eroded or blunted by heat
Heat-induced fractures can be aligned more properly during reconstruction
Table 1: Some major features of traumatic and heat-induced fractures as well as coexistence of both, as observed in former studies. Note similarities between traumatic and heat-induced fracture features.
5 Blunt force trauma (BFT), on which lies the focus of this study, is one of the most common trauma type and manner of death, worldwide [10,11,12]. Concentrated, dynamic external stress to bone from BFT results in fractures [13]. The interplay of elasticity and rigidity provided by organic-inorganic constituents of bone with its viscoelasticity and anisotropy accounts for the biomechanical properties of the bone [2,12]. These properties are also dependant on intrinsic factors like shape, structure of bone and extrinsic factors like strength, force, strain and stress – latter two being paramount for bone biomechanics [2,12]. Strain refers to the change or deformation of bone in response to external force while stress refers to the force per unit area [13]. Along with abovementioned factors, the stress-strain relationship of bone determines the type of deformation and possible failure of the bone as described in Figure 1 [12,14]. BFT, unlike gunshot trauma, is caused by slow-loading force and causes multiple ways of bone fracturing [12,13]. This study focuses on complete fractures caused by combination of compressive and tensile, angulation forces from the BFT impact [12,15]. Although traumatic fractures from skeletal remains are seldom indicative of cause of death, they are suggestive of the manner of death – defence, accidents, homicide or suicide [16]. This knowledge allows forensic experts and investigators to refute/support certain hypotheses and, when combined with tactical information and other evidence, the investigation can be steered towards more logical directions [12].
A body subjected to extreme fire will undergo several modifications before only bones remain. Exposure to thermal stress also alters the bone structure (macro and micro) and composition [1]. Such major alterations include shrinkage, warping, weight loss and deformation, fracturing, colour and dimensional changes [16]. Generally, bones burn through 4 stages namely dehydration (breaking of hydroxyl bonds with removal of water), decomposition (pyrolysis of organic constituents), inversion (loss of carbonates and crystals conversion) and fusion (melting and coalescing of inorganic crystals) [9,16]. During this process, the colour of bone changes from ivory to dark-brown (around 300°C) then carbonization of organic constituents causes black colouration (around 400°C) with their loss (above 500°C) causing calcination, seen as grey then white [16,17]. Thompson (2005) indicated that dimensional changes occur during the fusion stage. Moreover, the collagen dehydration reduces bone elasticity, resulting in deformation of bones, typically as different types of HIFs (Figure 2). These heat-induced fractures (HIFs) occur from evenly disseminated, static stress within bone [16]. Distinguishing HIFs from BFT-fractures allows proper hypotheses formulation about a case such as an accidental fire versus concealment of homicide [1].
Figure 1: The stress-strain curve. In bones, firstly, elastic deformation happens when bone resumes its original shape after stress is gone. Secondly, plastic deformation occurs when bone becomes irreversibly deformed even after stress is removed. Lastly, when the force surpasses the bone strength, failure occurs, causing a fracture (taken from [13]).
6 Extensive literature on effect of fire on bones and BFT as individual concepts exists. However, there is fewer literature discussing the effect of fire on bones with traumatic fractures [14,18,19,20] and only scarce studies dedicated solely to distinguishing BFT fractures from HIFs exist [3,7,11]. Situational fractures are also important fractures arising from burning bones but are not directly caused by heat. These fractures occur during late phases of burning and result from post-fire recovery and/or physical forces acting on the bones during such phases. It is important that these fractures are not confused with HIFs, so these will be evaluated as well. Unfortunately, most of these latter studies utilize defleshed animal models. Albeit being analogous to human bones, mineral density, hardness and microstructure of animal bones ultimately differ from human bones [1]. Hence, fracture propagation differs between human and animal bones, making translation of results from animal studies for actual casework involving human bodies infeasible [13]. To overcome this issue, the proposed project will be using human cadaveric bones, adding forensic value to the results from this project. Moreover, prior studies explored these features either individually (only colour changes, microscopic changes or dimensional changes) or in small groups (chiefly fragmentation, fracture surface, outline and acute/right/obtuse angles) ([16,17] about colour changes and [9] about dimensional changes). They also employ unstandardized and/or ambiguous descriptions for features such as ‘slightly transverse’ or ‘roughly V-shaped’ or ‘right angle’– no proper explanation for ‘slightly or roughly’ and if the ‘right angle’ must exactly be 90 or there is a range [11]. This project aims to combine and explore many of the of HIFs and BFT fractures features in a single checklist, with better-specified descriptions and explore these variety of features for eventual fracture type differentiation. Accordingly, the following research questions were investigated:
What features of the fractures are characteristic of BFT-fractures and HIFs (and situational fractures)? Which features of both types of the fractures overlap?
How prevalent are these characteristic and overlapping features in the bones?
How similar are the observed features to those features mentioned in existing literature? Is the checklist effective for the differentiation of fractures?
Is there a correlation between temperature, discolouration of bone and formation of heat-induced fractures? Is stereomicroscope applicable to effectively differentiating BFT-fractures from HIFs?
The strength and failure point differ for every bone type and depends on type of external force [13]. Similarly, the burning duration and pattern differ for varying bones types [1]. As such, this project focuses on only radius and ulna (long) bones, blunt force trauma (BFT) as the external force and calcination for the burning phase, to answer the above questions. BFT to the forearms, especially from self-defence, and calcined remains [1] are very common in forensic
Figure 2: Different types of HIFs- a) longitudinal, b) curved transverse, c) straight transverse, d) patina, e) delamination (Mayne Correia, 1990).
7 contexts.Fresh-frozen, defleshed forearm bones were grouped and subjected to only burning, only BFT, or both burning and BFT to identify only HIFs, only BFT-fractures and a combination of both types, respectively. The resulting fractures were examined both macro- and microscopically (stereomicroscope) using checklist (using both morphological and metrical observations) derived from previous acclaimed studies (Table 2). This checklist compiles features recorded by other authors and some new features to determine if these features can effectively aid the analysis of burned bones for prior BFT. Former studies only describe usage of expensive and relatively complicated analytical techniques such as SEM [3] or use only macroscopic analysis [11]. Thus, this study also aims to determine the usability of a stereo microscope, a much simple technique, in analysing microscopic traits of the different fractures. Stereomicroscope has been successful for a similar purpose -to scrutinize cut-marks in bones [1,21]. As a sub-question, the effect of maceration of fractures was investigated. Results from the entire study were statistically analysed and later compared with features and observations obtained by former studies, to determine consensus and discrepancies.
Feature of fracture(s) Studies which focused on respective features
Degree/State of burning of bone Mayne Correia, 1997; Herrmann & Bennett, 1999; Krap, van de Goot, Oostra, Duijst & Waters-Rist, 2017; Macoveciuc, Márquez-Grant, Horsfall & Zioupos, 2017 Fragmentation Herrmann & Bennett, 1999; Macoveciuc, Márquez-Grant, Horsfall & Zioupos, 2017 Colour of bone/fragments Mayne Correia, 1997; Herrmann & Bennett, 1999
Fracture category Macoveciuc, Márquez-Grant, Horsfall & Zioupos, 2017
Location of fracture on bone Mayne Correia, 1997; Herrmann & Bennett, 1999; Macoveciuc, Márquez-Grant, Horsfall & Zioupos, 2017
Fracture outline, surface morphology and angle
Villa & Mahieu, 1991; Mayne Correia, 1997; Herrmann & Bennett, 1999; Outram, 2004; Wieberg & Wescott, 2008; Wheatley, 2008; Poppa et al., 2011; Macoveciuc, Márquez-Grant, Horsfall & Zioupos, 2017
Dimensions of fracture and bone Thompson, 2005; Waltenberger & Schutkowski, 2017
Table 2:The various features employed in current study and the corresponding former studies, which investigated fractures using these features.
8
3. MATERIALS AND METHODS
The study was executed in two parts – a pilot study to determine ideal conditions for producing BFT-fractures in forearm bones and for burning, and a main study to explore the varying fracture features. Nonetheless, certain factors remained constant for both parts, such as the sample material, the custom-made rod-based contraption, the checklist of features (Appendix, Figure 1) and the method of fracture visualization and examination of bones. The sample preparation and experimental setups for the two parts have been explained below.
3.1. Sample material 3.1.1. Sample preparation
Fresh-frozen human cadaveric forearm material from the body donation program of the Department of Anatomy, Embryology and Physiology of the Amsterdam Universitair Medische Centra (UMC), The Netherlands, was used. In total, 38 bones (19 radius and 19 ulna) were obtained, thereafter manually defleshed with a scalpel to ensure that there was no added stress on the bones, producing additional fractures and were stored at 4-8°C, throughout the course of the study. Population data (age, sex and bone dimensions) was also collected for each of the 38 fresh-frozen human forearm bones (Appendix, Table 2). There were 28 bones from females and 10 bones from males. The age range spans from 64 to 88, with most cadavers being more than 75 years of age. The bones used for the pilot study (shaded rows in Appendix, Table 2) weighed the least and other dimensions were also smaller compared to the remaining pool of 30 bones, which were used for the main study.
3.1.2. Confounding factors
Since age, sex and medical history of the cadavers are confounding factors [11], cadavers were carefully selected – older aged individuals, equal number of males and females and no osteoporosis, bone cancer, surgery or other bone defects.
3.2. Pilot Study
This part focused on determining the optimal conditions factors and conditions for (i) generating the BFT-fractures and (ii) burning of the bones. The research questions for this part were:
What are the ideal experimental conditions for:
(i) Creating intended BFT-fractures (impact force and angle of this force) and
(ii) Effective calcination of fresh-frozen forearm bones in pyre (temperature and duration of burning)? 3.2.1. Pilot study (i): BFT generation
BFT was created on 5 fresh-frozen, defleshed bones by the custom-made contraption, resembling a pendulum apparatus (Figure 3). The rod attached to cylindrical impactor mimics the common weapons involved in BFT cases such as baseball bats. A grading element (0 to 180) indicates the angle at which the rod is released from. To determine the ideal angle needed to produce the intended fractures, the rod was released from various angles till the bones showed similar fractures. The impact area of the rod on the bone was visualized using Pelikan green ink – label sticker was
9 placed around the cylindrical impactor and dabbed with the ink. The rod was slowly released onto the bone till the ink transferred to the bone, representing the impact area. The impact energy, area and force needed to generate the fractures were then calculated using formulae presented in Appendix (Box 3). Forearms were positioned on the contraption in a constant manner to standardize fracture production – thicker side of bone facing the impact rod, middle of diaphysis touching the rod and head of radius/olecranon of ulna placed on the right holder (Figure 4). The rod was held at the middle of the longitudinal axis of the rod and at 80,before being released.
3.2.2. Pilot study (ii): Burning temperature and duration
2-5 cm transverse sections of the diaphysis of 2 radial bones were made using hand-saw, whereby the bone was kept wet to avoid undesirable heat from the friction of sawing. These sections were placed in small porcelain cups were heated in a muffle oven (accuracy of 2C). A temperature range of 600°C to 700°C and time intervals of 25 to 45 minutes were utilized to determine ideal temperature and duration for attaining the inversion to calcination stage of burning. Owing to the great difficulty of controlling fire temperature to accurately stop the burning at any prior stages and collect bones within stipulated time limits, this burning stage has been selected [1].
The obtained fracture-production and burning conditions were used as the standardized conditions for the main study. Furthermore, the checklist was tested in this part to ensure the chosen list of features were appropriate and feasible for the remainder of the project.
Figure 3: Custom-made pendulum-like contraption to produce fractures. The rod is attached to a cylindrical impactor (red arrows), together weighing 3kg. A grading element (black arrows), ranging from (0 to 180) shows the angle at which the rod is released from.
Figure 4: Placement of bones on the custom-made pendulum-like contraption to produce fractures. Yellow arrow shows the right side of the device, where the olecranon and radial heads were placed – olecranon facing the rod and radial tuberosity facing away from the rod.
10 3.3. Main study
This part focused on identifying the fracture characteristics. Forearm bones were split into 3 groups (Table 3; Appendix, Table 4 to see which bone specimens are in which group) with 10 bones in each group (equal number of radius and ulna bones in each group). with the sex, type of bone (radius/ulna) and dimensions being distributed as evenly as possible across the groups. This distribution of parameters in every group is depicted in the box plots (Figure 5) and Table 4. The medians of each group, for age and length, are quite similar while Group B shows larger median than rest, for weight. There is not much variation in the minimal-maximal values for age and length parameter across the groups. Groups A and B have greater maximum for weight than group C. These variations arise from usage of a limited sample size with desired criteria (preventing confounding factors). There were also only 4 out of 30 bones weighed above 100g and had to be split as evenly as possible amongst the groups. The data are all skewed towards the upper quartile for age, weight and length. Each group were subjected to standard fracture-production and burning conditions, obtained from the pilot study, wherever necessary.
Group name Type of exposure Specimen label A Only burning & no BFT (control) RAX or UAX B Only BFT & no burning RBX or UBX
C Both burning and BFT R/UCX (PrB) and R/UCX (PoB)
Table 3: Grouping of forearm bones, each group with equal number of ulna and radius. Type of exposure indicates the type of testing each group of 10 bones were subjected to. Abbreviation for specimen labels: R- radius, U- ulna, A/B/C- group name, X- number of bones in that group (1 to 10), PrB- Pre-burning and PoB- post-burning.
Sex Group A Group B Group C Number of radius and ulna Group A Group B Group C
Male 5 5 6 Radius 5 4 5
Female 5 5 4 Ulna 5 6 5
Table 4: Indicates the distribution of bones from males and females (right) and radius/ulna (left) in every group. Due to inequality in the total number of bones from males (16) and total number of bones from females (14) for the main study, Group C has more bones from males than females. Due to inequality in the total number of radii (14) and total number of ulna (16) for the main study, Group B has more ulna than radii.
Figure 5: Box plot showing the distribution of bone samples in each study group, based on age (left), weight (center) and length (right) of respective cadavers. Each group mostly has bones from adults ranging between 70-84 years old. Each group mostly has bones weighing between 70-85g. Groups A and B each contain one of the 2 heaviest bones (in all 30 bones). Every group mostly has bones ranging between 25-27cm. Each group also contains one of the 3 shortest bones (in all 30 bones).
11 3.3.1. BFT production
All bones were struck using the pendulum-like contraption, as detailed in 3.2.1. 3.3.2. Maceration
The maceration for Group B was done by placing the 10 bones in 100%-nylon socks (to collect the fragments) and cooking them only in water (till 100C; no soap or chemicals) for 26 hours (Figure 6). The bones were then lightly scrubbed using a sponge and left to cool at room temperature for 24 hours before further analysis.
3.3.3. Burning
The bones were burnt in Fire Station Soest, in a room of an isolated concrete firehouse, which resembled a basic residential living room (mimicking forensic cases) (Figure 7). The experiment was conducted on a sunny day with mild wind. The pyre consisted of mixture of hardwood (±8kg)-softwood (±3.5kg) under two (120cm X 80cm) wood pallets. A foam (polyurethane) mattress covered with cotton mattress-cover (70%-cotton) was placed on the pallet. The two groups of bones (A and C) were encased into ‘packages’, using mesh-wire (hexagonal-shaped, 13mm perimeter), 100%-cotton shirts and pig skin (15kg and approximately 1.33cm of adipose tissue as fuel). The entire set-up (Figure 8) was ignited using 4 wood-blocks and lighter, no accelerant was used. The windows and doors of the firehouse were opened to allow oxygen supply to the fire. The bones were burned for 50 minutes, whereby the fire was stable, until the last 5 minutes when it was contained to each bone-package. The approximated temperature of the fire ranged between 700C - 800C, based on temperatures reached by fuel materials (mattress, wood and pig skin). The fire was put out by a fireman with water spray (3 minutes of spraying). The bones were carefully picked out from the burnt pyre/package and placed in buckets for transportation to the institute. The residual water from the bones was removed by carefully blotting the bones with tissue paper. The bones were left to cool and dry for 24 hours before further analysis.
Figure 6: Maceration set-up is shown here. Group B bones were carefully placed into grey and black coloured nylon socks, which were knotted and kept into a wire cage. The cage was submerged into a container of water, fully covering the top of the cage). The water was slowly heated to 100ºC and bones were macerated, without any chemicals.
Figure 7: Schematic representation of the room in firehouse that was used for burning the bones. Note furniture and placement of wooden pyre
12 3.4. Observations: Macroscopic and microscopic analyses
Fractures were analysed according to the checklist derived from former studies (Appendix, Figures 1-2). Macroscopic and microscopic observations of these bone fractures were made according to the checklist. The prevalence of features was evaluated, intra-group and inter-group (Table 5).
Compared groups Outcome of comparison
Group A with Group B Identification of specific features of HIFs and BFT-fractures, respectively Group A with Group C Determination of HIFs and other overlapping features
Group B with Group C Determination of BFT-fractures and other overlapping features
Table 5: Intergroup comparisons and the different outcomes in relation to specifying features of each fracture type.
Figure 8: Burning experiment set-up is shown here. A:Bone packages stacked on either side of mattress – Group A as Group I at left and Group C as Group II at right; B: 2 layers of wire mesh layered and laid out in criss-cross manner, on top of wooden pallet, wood and mattress; C and D:Packaging process of bones – shirt unbuttoned and flattened on top of wire meshes, pig skin evenly spread on shirt, bones arranged on pig skin, pig skin wrapped above bones, shirt closed over pig skin and buttoned up before wire meshes are tightened and tied together. E: Mix of hardwood and softwood, depicted by cyan arrows, placed below wooden pallet within the firehouse; F: Placement of packages and mattress on wood-wood pallet set-up; Red arrows: Indicate positions of wooden blocks for fire ignition.
13 3.4.1. Fracture visualization
All bones were photographically documented after each phase of experiments. Microscopic observations were made using the Olympus SXZ9 stereomicroscope, under 8X to 10X. This enables quick and easy microscopic analysis of the fractures, which aids in scrutinizing specific features of these fractures such as angle and morphology of fracture [21]. No preparation of the burned bones was done, allowing their usage for additional/future analyses/studies. Only one group (only-BFT) was subjected to maceration for better fracture visualization and to identify any differences between fracture traits of pre-/post-maceration of bones. ImageJ was utilized to further analyse the features and determine angle of fractures. The dimensions of the bone were measured as length of fracture on proximal/distal bone and ratio of length on tension side to compression side (Appendix, Glossary).
3.4.2. Measurement of angles
For the angle of fracture, apart from microscopic observation, the reconstructed bones were photographed and a negative-slope (downhill) was drawn across the proximal end of the fracture, spanning from the compression to tension side of the bone. Thereafter, the angle was measured as in Figure 9, thrice (blindly) at different times, to account for observational errors and the mean angle for each bone was determined. The standard error of measurement was calculated for each bone and the angle of fracture.
3.4.3. Colourimetric analysis
The colour changes were assessed using mirror-reflex camera and an unpublished colourimetric model developed by Krap et al [17]– the L (lightness) and B-coordinate, in form of cluster data, were used to correlate the colour of bones (in that study) to specific temperature ranges that the bones were subjected to. This model has demonstrated 86% accuracy. Photographs of the burned bones from this study were taken using a Nikon D700 with Nikon 35mm AF-D F2.0 and 24mm AF-D 2.8 lenses, then adjusted for major white balance and converted to L*a*b colour space (ImageJ – colour transformer plugin) to determine the L (lightness) and B-coordinate. For each bone, 4 sections (Figure 10) were manually selected to calculate the average for temperature range comparison (data from unpublished model). Measurements were made slightly away from the margins of the bone to avoid influence of reflectance from bone, on the L-b values. The comparison of these values (corrected ones) to the model allows determination of the temperature range to which that region of bone was exposed. For calibration of the colourimetry, X-rite’s ColorChecker Classic target was used – one photo before photographing the burned bones and one after, then averaged. The difference between the manufacturer values of L-b and adjusted L-b values was subtracted from the L-b values measured from the photographs of bones. Due to brittleness of bones, the anterior portion (mostly) of the bones was photographed.
Figure 9: Angle measurement using ImageJ – downward/negative/downhill line drawn from compression to tension and angle (red) determined. Left: Bone subjected to BFT; Middle: Macerated bone; Right: Burned bone. Images are of 3 different bones.
14 3.5. Statistical analysis
Only the results from the main study were statistically evaluated. Table 6 shows the dependent and independent variables associated with this study. MS-Excel and IBM SPSS Statistics 25 were utilized for statistical analysis. Descriptive statistics was done for all groups. Since the fracture features correspond to categorical (colour, stage of burning, fracture outline, etc.) and numerical data (dimensional changes), Kruskal-Wallis test and one-way ANOVA analysis were done respectively to identify statistical significance of difference in features (such as fracture outline, surface, types, burning phases or dimensions) between the two compared groups [22]. These analyses enable the identification of BFT/HIF-specific features and/or any overlapping features – both key research questions. Due to small sample size (n=10 per group), Q-Q plots were used to determine normality of data and Levene’s test for homogeneity of variance was also done; both being requirements for one-way ANOVA analysis. The categorical data were numerically-labelled/scored to aid in this analysis, as in Table 7. The group names (A, B, C) were used as the ‘factor’ and each feature as the ‘response’. Owing to the small sample size, a significance level of 0.05 was used [23]. Post-hoc analysis was not done as only 2 groups were compared at a time.
Finally, these features were cross-compared with current literature, in terms of feature description and occurrence in each fracture type, to assess and explain similar and conflicting results.
Dependent Independent
Type of HIF Age range and distribution of sex of human cadavers Features of HIFs that mimic/are similar to features of
BFT-fractures
Type of bones used - radius and ulna
Features distinguishing HIFs from BFT-fractures Site of BFT impact on bone – middle of diaphysis and positioning of bone on contraption (hand pronation) Force, velocity, mass and angle of BFT impact on bone Duration and temperature of burning
Type of burning environment
Table 6: Dependent and independent variables associated with the study.
Feature Corresponding numerical label/score Fracture category
Complete-simple 1
Complete-comminuted 2
Incomplete 3
Fracture outline Helical/ curved 1
Transverse 2
Figure 10: Sections used to measure L-b values –1/4 proximal, 2/4 proximal (Fx proximal in BFT bones), 3/4 distal (Fx distal in BFT bones) and 4/4 distal. Note how each section is marked (red and yellow) slightly away from cortical bone margins.
15
Longitudinal and transverse 3
Diagonal 4
Diagonal with a step 5
Columnar 6 Fracture location Proximal 1 Intermediate 2 Distal 3 Fracture surface Smooth 1 Rough 2
Rough and smooth 2
Fracture type Transverse 1 Oblique 2 Spiral 3 Comminuted 4 Segmental 5 Longitudinal 6 State of burning Unmodified 1 Carbonized - Early 2 Carbonized - Complete 3 Partially burnt 4 Calcined - Partial 5 Calcined - Complete 6 Complete 7 Colour of bone Black 1 Dark grey 2 Light grey 3
Grey (balance of light & dark) 4
Blue-Grey 5 White 6 Brown 7 Type of HIF Longitudinal 1 Straight transverse 2 Curved transverse 3 Step 4 Patina 5 Delamination 6 Warping 7
Temperature distribution (from colourimetry)
300C -600C 1
450C -700C 2
700C -900C 3
> 900C 4
16
4. RESULTS
4.1. Pilot study
4.1.1. (i): BFT generation
Table 8 shows the different angles used to generate fractures on the bones. Apart from 40 and 50, which failed to produce fractures, the other angles resulted in complete fractures. The ideal angle was selected based on whether the impact will be consistent among all sample bones, despite the varying bone densities. At 130, the impact was deemed to be too strong to be used for bones with lower densities. At 60, the impact was deemed to be insufficient for bones with higher densities. As such, 80 was chosen to allow consistent impact across all bones. The area of impact on bone (depicted in 14917U-Lt of Table 8) and the impact force were calculated to be 1.65 × 10 𝑚 and 2066𝑁 − 4650𝑁 respectively (calculations provided in Appendix, box 3).
Specimen
Angle at which rod is released
from ()
Observations Images
04018U-Lt 130 Complete fracture with tiny
fragments
04018R-Rt 90 Complete fracture
04018U-Rt 70 Complete fracture
10817R-Lt 40 No fracture
10817R-Lt 50 No fracture
10817R-Lt 60 Complete fracture
14917U-Lt 80 Complete fracture
Table 8: Observations from the pilot study to identify the ideal angle for the generation of BFT fracture, which is denoted by the yellow highlight. 14917U-Lt shows the impact area (an ellipse) caused by the rod while fracturing the bone.
17 4.1.2. (ii): Burning temperature and duration
Table 9 indicates the results obtained for determining the ideal temperature and duration for the transitionary burning stage from inversion to calcination. The temperature and duration were varied by trial-and-error method and depending on the result observed in the preceding bone-section. The highlighted sections show the most apt temperature and duration for the preferred burning stage, which can be collated as: 670C to 690C for temperature and 30 to 40 minutes for duration. One bone section with the epiphysis extending into the diaphysis was also utilized to ensure if the obtained result could be replicated in a longer section of bone with slightly more soft tissue, which was successful. The burned bones were also scrutinized as per the formulated checklist. The checklist was indeed effective, whereby many features (except microscopic observations) were characterizable according to the list. Few heat-induced fractures were also seen. Specimen Length (cm) Temperature (°C) Duration of burning (mins) Observations Images
10817R-Rt 4.7 600 25 Dark grey and some grey-blue colouration with longitudinal HIF
10817R-Rt 3.9 620 30
Mostly grey and grey-blue with some yellowish-orange regions, early longitudinal HIF
04018R-Lt 2.2 650 25
Mostly grey and grey-blue with patches of yellowish-orange and white regions, no HIF; medullary cavity more white than previous burned specimens
18
04018R-Lt 2.1 650 35
Almost 50-60% white only on one side of bone section, less grey but more grey-blue areas, spongy layer more clumped together, no HIF
04018R-Lt 2.3 670 40
Almost fully white with slight tinge of light grey, sawn edges are white, spongy layer shows separation from cortical bone, no HIF
04018R-Lt 2.1 690 25
Largely grey/grey-blue with patches of white, slight cracking of bone seen
04018R-Lt Epiphysis
5.6 690 35
95% white with some grey areas and spots of yellowish-orange; longitudinal and curved transverse HIFs seen
10817R-Rt 4.1 680 30 Mostly white with tinge of grey, sawn edge is white, no HIF
Table 9: Observations from the pilot study to identify the ideal temperature and duration of burning for the inversion to calcination stage to occur. Yellow highlighted sections denote the afore-mentioned factors apt for the desired stage of burning.
4.2. Main study
4.2.1. Post-BFT features (Pre-Maceration and Pre-Burning)
Bones from Group B (Pre-Maceration) and Group C (Pre-Burning) mostly (15/20) showed fractures on the fractures intermediate part of the bone (Appendix, Tables 11-12). Except three bones, the remaining bones had only one fracture. 60% of bones showed fragmentation and, of varying sizes. There were more complete-comminuted fractures (11/20) than completed-simple fractures (9/20), in terms of fracture category (Appendix, Figures 6-7). All bones showed a smooth fracture surface morphology. Helical/curved was the most common fracture outline, followed
19 by, transverse, diagonal, columnar and longitudinal-transverse. 12 out of 19 bones (excluding RC2) were oblique fractures, followed by, spiral, transverse, comminuted and one segmental fracture. Group B resulted in greater fracture lengths (predominantly 10-15cm) on the proximal side (proximal: 90%, distal:70%) than the distal side of the bone and vice versa for Group C (proximal: 55.6%, distal 77.8%). The ratios of fracture length of tension to compression side was greater on the proximal side than distal side for Group B (proximal: 90%, distal:70%) and the same for Group C (both 55.6%). Typically, these proximal-distal ratios were nearly the same for bones with transverse fractures (UB7 and UB9). Only RC7 showed a segmental fracture and was not statistically evaluated to avoid large deviations in fracture types. Q-Q plots exhibited normality for the numerical features (length, ratio and angles of fractures) while Levene’s test indicated homogeneity of variances. ANOVA analysis of the numerical features between Groups B and C did not show any statistical significance (p-value 0.05, all means were equal for inter-group) except for the angle of fractures (Table 10). KW-test also showed no statistical difference for the categorical features between the groups except for type of fracture and location on bone.
ANOVA – Feature df Sig. Fx Length, Proximal bone 1 0.986 Fx Length, Distal bone 1 0.983 Ratio of length, Proximal bone 1 0.872 Ratio of length, Distal bone 1 0.842 Fx angle 1 0.007 KW test – Feature Fx surface Fx outline Type of Fx Fx category Location on bone No. of fragments Fragment size df 1 1 1 1 1 1 1 Asymp. Sig. 1.000 0.206 0.042 0.849 0.039 0.719 1.000
Table 10: ANOVA (above) and Kruskal Wallis test (below) results for post-BFT – Group B vs Group C. Yellow highlight indicates statistically significant result.
4.2.2. Post-Maceration (Group B)
The macerated bones were slightly greyish in colour due to mild staining from the nylon socks (black and grey-coloured) but this did not mask any visual characteristics of the bone. The fracture lines were clearer and more defined in macerated bones than unmacerated ones due to lack of any soft tissue (Appendix, Figure 6 and Table 13). However, some macerated bones had more fragmentation after maceration, especially at the previously fractured and/or fragmented sites (see UB2, RB3 of Appendix, Figure 6). This fragmentation, along with extending spongy layers (longer than cortical bone at fracture site) led to reconstruction of the bones being more difficult. The sharpness of fracture edges was almost completely lost in 3 macerated bones and 6 others showed more bluntness with one/two sharp shards while only one macerated bone retained its sharpness (Figure 11). The fracture category remained unchanged, pre- and post-maceration, except for UB2 that showed fragmentation post-maceration. All bones showed a smooth fracture surface morphology. Transverse fracture outline was the most common, followed by diagonal and helical/curved. Except for UB4, there were no differences in fracture type between unmacerated and macerated bones. There were only some minor changes (maximum 0.4cm) in the proximal and distal lengths of the macerated bones as compared to unmacerated bones. The same applies for the ratios of fracture length of tension to compression side. Normality and Levene’s test were satisfied for the numerical features and ensuing ANOVA analysis showed no
20 statistical difference between unmacerated and macerated bones (Table 11). The same applies for KW-test of categorical features between each group except for sharpness.
ANOVA – Feature df Sig. Fx Length, Proximal bone 1 0.727 Fx Length, Distal bone 1 0.626 Ratio of length, Proximal bone 1 0.352 Ratio of length, Distal bone 1 0.723 Fx angle 1 0.871 KW test – Feature Fx surface Fx outline Type of Fx Fx category Location on bone No. of fragments Fragment size Sharpness df 1 1 1 1 1 1 1 1 Asymp. Sig. 1.000 0.677 0.689 0.648 0.168 0.167 0.684 0.030 Table 11: ANOVA (above) and Kruskal Wallis test (below) results for Group B – unmacerated vs macerated.
4.2.3. Post-Burning (Group A and C)
Group A bones were tough to be individualized and were evaluated as a group with new specimen/sample numbers (Appendix, Figure 5 and Table 14). Group C bones were individualized to original specimen numbers thus being easily comparable to previous results (Appendix, Figure 7 and Table 15). All bones were very brittle (Group C bones more brittle than Group A), thus resulting in fragmentation during post-fire recovery (some fragments were missing too), transportation to the laboratory and subsequent macro/micro analysis. Hence, fragmentation but was not statistically evaluated. 16 out of 20 bones showed large areas of partial calcination and small areas of carbonization, the remaining bones were predominantly carbonized with small regions of partial calcination. Group A bones were tough to be individualized and were evaluated as a group with new specimen/sample numbers. Group C bones were individualized to original specimen numbers thus being easily comparable to previous results. Moreover, as RC7 was excluded for evaluation in 4.2.1, it was not examined here too. Both groups showed fragmentation but was not statistically evaluated since some fragments were missing during post-fire recovery or further broken during transport to laboratory. The discolouration and state of burning of both groups did not show any distinct trend although it seems that proximal ends were more carbonized than rest of bone and calcination begins from the intermediate portion of the
Figure 11: Example of microscopic observation of sharpness pre-maceration (A1 and B1) and post-maceration (A2 and B2) for Group B bones. The yellow oval-markings indicate the loss of sharpness of edges (A1 to A2). The purple oval-markings demonstrate the unaltered region of the one bone which retained its sharpness. Magnification: 8X.
21 bone. As before, Group C bones mostly showed helical/curved and diagonal fracture outlines while Group A bones showed transverse fracture outlines (only 5 showed situational fractures). Both groups exhibited clearly defined, blunt fracture edges but Group A had smooth surface whereas Group C largely had rough surface and some smoothness in curved/sloped regions of fracture edges (Figure 12). Longitudinal HIFs followed by straight transverse and step HIFs were prominent in both groups while situational fractures were predominantly longitudinal. Situational fractures displayed fresh margins and edges with even spread of rich colour, but BFT-fractures show slight unevenness of colour at edges (Figure 14). Interestingly, fracture lines of HIF penetrated into the medullary cavity, but fracture lines of situational fractures were elevated on one side (Figure 13). Heat border was also observed close to BFT site of Group C bones, denoting transition of burning phases of bone. Microstructural fractures were found in partially calcined bones with whiter colouration. Bones with greater calcination also showed better articulation than bones with greater carbonization. Normality and Levene’s test were fulfilled for the numerical features and ensuing ANOVA analysis showed no statistical difference between both groups (Table 12). KW-test resulted in statistical difference for fracture surface, outline, category and location on bone while other features had no significance. Comparing unburned Group C bones with burned ones, ANOVA and KW-test (Table 13; Appendix, Tables 13 and 16) resulted in statistical difference only for fracture surface and sharpness. Most bones had lost their sharpness. As for bone dimensions, most bones in Group C exhibit shrinkage, with some increases in proximal length. Only one bone (UC10), however, shows increased lengths for both proximal and distal part of the bone, denoting an expansion across the bone. The ratios of length on tension to length on compression for proximal and distal bone have mostly decreased, except for few increases on proximal side and one on distal side.
Figure 12: Depicts the presence of both roughness and some smoothness (at sloped/curved edges) in burned bone from Group C. Purple arrow indicates the irregular, rough region and yellow arrow indicates the relatively smoother region.
22
A1 B1
A2 B2
Figure 13: Depicts the same plane of margins for HIFs and elevation of margin on one side of situational fracture. Top, left: Longitudinal HIF; Top, right: Longitudinal situational fracture where margins on top part of fracture is elevated. Middle, left: Curved longitudinal situational fracture running through BFT site and surrounded by microstructural cracking; Middle, right: Note how the situational fracture has margins on an uneven plane, as compared to surrounding cracking. Bottom, left: Schematic eye-level representation of elevated margins in situational fractures (A1) vs margins on same plane as longitudinal axis of bone in HIFs (B1) and pink-shading depicts spongy layer; Bottom, right: Schematic transverse view of margins in situational fractures (A2) vs HIFs (B2). In bottom images, green line indicates longitudinal axis and brown line shows transverse axis.
Figure 14: Denotes the difference in margins appearance and colouration of edges for situational fracture (A1 and A2) and BFT-fracture (B1 and B2). A1 shows the even spread of distinctive grey discolouration of BFT-fracture edges while A2 shows the fresh margins. B1 and B2 show heat-altered margins and edges with uneven discolouration, as represented by the blue and red arrows – each coloured arrow points at two different colours in each image.
23 ANOVA – Feature df Sig. Fx angle 1 0.658 KW test – Feature Burning state Top colour Second common colour Third common colour Fx surface Fx outline Fx category Location on bone Most common fire Second common fire Not directly fire df 1 1 1 1 1 1 1 1 1 1 1 Asymp. Sig. 0.206 0.450 0.529 0.735 0.000 0.038 0.003 0.007 0.051 0.962 0.322 Table 12:ANOVA (above) and Kruskal Wallis test (below) results for post-burning – Group A vs Group C. Yellow highlight indicates statistically significant result.
ANOVA – Feature df Sig. Fx Length, Proximal bone 1 0.864 Fx Length, Distal bone 1 0.880 Ratio of length, Proximal bone 1 0.587 Ratio of length, Distal bone 1 0.977 Fx angle 1 0.280 KW test – Feature Fx surface Fx outline Type of Fx Fx category Location on bone Sharpness df 1 1 1 1 1 1 Asymp. Sig. 0.000 0.775 0.469 0.331 1.000 0.000
Table 13: ANOVA (above) and Kruskal Wallis test (below) results for Group C – pre-burning vs post-burning. Yellow highlight indicates statistically significant result.
4.2.4. Comparison of BFT and HIF features between groups
Normality and Levene’s test were satisfied for the statistical analyses described hereafter. ANOVA and KW-test of Group A-Group B indicated statistical difference for fracture angle, outline, category, location on bone and fracture type (Table 14). They did not have any significant difference for the fracture surface, number and size of fragments. Only fracture surface was similar between these two groups. Comparison of Group B and Group C showed statistical significance for fracture angle, length of fracture for distal bone, fracture surface and type of fracture obtained (Table 15). These groups did not show significant differences for fracture outline, category, proximal length and ratiosn for both sides of the bone.
ANOVA – Feature df Sig. Fx angle 1 0.038 KW test – Feature Fx surface Fx outline Type of Fx Fx category Location on bone No. of fragments Fragment size df 1 1 1 1 1 1 1 Asymp. Sig. 1.000 0.026 0.000 0.009 0.049 0.076 0.111
Table 14:ANOVA (above) and Kruskal Wallis test (below) results for post-burning Group A vs BFT only Group B. Yellow highlight indicates statistically significant result.
24 ANOVA – Feature df Sig. Fx Length, Proximal bone 1 0.181 Fx Length, Distal bone 1 0.027 Ratio of length, Proximal bone 1 0.245 Ratio of length, Distal bone 1 0.667 Fx angle 1 0.009 KW test – Feature Fx surface Fx outline Type of Fx Fx category Location on bone df 1 1 1 1 1 Asymp. Sig. 0.000 0.272 0.025 0.265 0.051
Table 15:ANOVA (above) and Kruskal Wallis test (below) results for BFT only Group B vs post-burning Group C. Yellow highlight indicates statistically significant result.
4.2.5. Colourimetric analysis
The two groups of bones, arranged on 4 white sheets of paper were photographed from top-view and visually, most bones show partial calcination (Figure 15). Unfortunately, only anterior portions of most bones were photographed. Upon comparing the L-b values to the unpublished model, 14/20 bones, from Group A and C combined, were observed to have undergone temperature of 450C -700C (Appendix, Tables 8 and 9). Only 4/20 bones demonstrate regions corresponding to temperature of 700C-900C. Group C bones display more variation in burning pattern across the bones (Figure 16) than Group A bones, which show greater similarity (Figure 17). Majority of bones from Group A converge at a temperature of 450C -700C, at the 2/4th portion of the bone (proximal side of the
diaphysis). Contrarily, Group C bones converge at a temperature of 450C -700C, at the distal side of the fracture site. There seems to be more carbonization at the epiphyses of most bones, than calcination. Contrarily, some bones are more calcined throughout the bones and less charred. There is no distinctive trend seen when comparing the location of HIFs and situational fractures on the bones with the burning pattern/discolouration. KW-test of temperature distribution (Table 16) showed significant difference at the 2/4-proximal region of the bone while other parameters did not show any statistical significance.
25 0 1 2 3 4 1 / 4 P R O X 2 / 4 P R O X 3 / 4 D I S T 4 / 4 D I S T TE M PE RA TU RE R AN G E PORTION OF BONE D I S T R I B U T I O N O F T E M P E R A T U R E R A N G E A C R O S S E A C H B O NE F O R G R O U P A 1U 2U 3R 4R 5R 6U 7U 8U 9R 10R 0 : < 300C 1 : 300-600C 2 : 450-700C 3 : 700-900C 4 : > 900C
Figure 17: Graph shows temperature ranges across each portion of bone for Group A, based on the unpublished colourimetric model. Most bones converge at 450C-700C, at the 2/4 proximal bone.
0 1 2 3 4 P R O X F X P R O X F X D I S T A L D I S T TE M PE RA TU RE R AN G E PORTION OF BONE D I S T R I B U T I O N O F T E M P E R A T U R E R A N G E A C R O S S E A C H B O NE F O R G R O U P C
UC1 RC2 RC3 UC4 RC5 UC6 RC7 UC8 RC9 UC10
0 : < 300C 1 : 300-600C 2 : 450-700C 3 : 700-900C 4 : > 900C
Figure 16: Graph shows temperature ranges across each portion of bone for Group C, based on the unpublished colourimetric model. Most bones converge at 450C-700C, at the Fx distal portion of the bone.
26 KW test – Feature 1/4 proximal 2/4 proximal 3/4 distal 4/4 distal df 1 1 1 1 Asymp. Sig. 0.260 0.031 0.490 0.376
Table 16: Kruskal Wallis test results for the temperature distribution (calculated using colourimetry) of Group A vs Group C (post-burning). Yellow highlight indicates statistically significant result.
4.2.6. Dimensional changes
Due to difficulty in individualizing burned bones in Group A, they were evaluated as a group and there was an average of 1.13% shrinkage. As for Group C, the dimensional changes from pre-burning to post-burning have been briefly discussed in 4.2.5. There was a maximum shrinkage of 4.61% and minimum of 0.75% for the proximal length (Mean: 2.63%) and maximum shrinkage of 19.31% and minimum of 1.64% for distal length (Mean: 8.44%). Most of the ratios of length on either side of the bones have reduced and are closer to 1.Some bones show expansion due to increased proximal and distal fracture lengths as well as increased ratios - maximum of 20.31% increase and minimum of 0.75% increase for fracture lengths and, maximum of 38.82% increase and minimum of 4.60% increase for the ratios. The shrinkage is more spread out for the distal fracture length, with seemingly greater shrinkage than expansion. This spread is much lesser for the proximal fracture length and the ratios. Figure 18 and Figure 19 depict the spread of percentage decrease (shrinkage) and percentage increase (expansion) in burned Group C bones, respectively. The lengths of fracture on the distal side and the ratios of length on the proximal side show more variation for both shrinkage and expansion. The means for each dimensional parameter also differs for both shrinkage and expansion. However, it is vital to note that only one bone showed increased ratio on the distal side and an outlier is present for the group of decreased ratios on the distal side.
Figure 18: Boxplot for shrinkage of bones Group C (post-burning) for each dimensional parameter. Length of fracture at distal portion of the bone and ratio at the proximal side show more variation while length of fracture at proximal side shows least variation. An outlier is seen for the ratio at the distal side.
27
Figure 19: Boxplot for expansion of bones Group C (post-burning) for each dimensional parameter. Larger variation of expansion percentages is seen for distal fracture length and the proximal ratio. Only one value exists for the distal ratio hence a single yellow line is depicted.
28
5. DISCUSSION
Concealment of crimes using fire is commonly encountered in forensic investigations. BFT is also frequently seen in forensics. Often, great difficulty exits in differentiating features of injuries such as fractures, caused by BFT from those caused by fire. This is particularly tough when both types of fractures co-exist [3]. The current study investigated the features distinguishing BFT-fractures from HIFs, by employing a novel checklist that collates different features described by prior studies. As discussed in the introduction, these studies rely on older definitions or use vague definitions that make fracture feature analysis tricky. Moreover, the features in such studies are descriptive/categorical rather than being metric/numerical, particularly colour and fracture angle. The checklist in this study strives to overcome these issues by using more specific definitions (Appendix- Glossary) and quantitative observations for key features. 5.1. Experimental design
It is crucial to comprehend and compare the experimental design of this study with other literature, so that there can be better understanding of the results and their relevance to application in forensic casework. The advantage and significance of using human material instead of animal material has been detailed in the introduction. The manner of storage and preservation of cadavers/bones also matters. Typical effects of freezing and embalming are water-recrystallization and formalin-caused collagen crosslinking, respectively [24,25]. Studies have shown the impact of such effects on bone biomechanics to be lesser in frozen bones than in embalmed bones [26, 27]. Usage of fresh-frozen stored up to 1year and with 1 freeze-thaw cycle is recommended [28,29]. As such, fresh-fresh-frozen bones with 1 freeze-thaw cycle were used.
As Ubelaker discussed, many antemortem and taphonomic factors such as age, body weight, previous illnesses and environment in which death occurred, influence the fracture propagation and corresponding features seen in bones. In view of this, the present study aimed at standardizing as many of these factors as possible, whilst maintaining coherence to forensic casework. A main factor would be the variability between humans in terms of bone dimensions, bone density, proportion of skin, fat and muscle [11]. The proportions will affect the fracture propagation and burning temperature of bone since these layers act as fuel. Thus, defleshing was done to allow more consistent fraction generation and burning temperature across bones subjected to fire. In forensic casework, it is uncommon to find macerated bones subjected to fire. Thus, use of unmacerated bones in this study is more relevant for actual cases. The lack of statistical difference between unmacerated and macerated bones allows comparison of results from the current study to those from previous studies, which utilized macerated bones. The minor differences of loss of sharpness and fragmentation between the two groups could lead to poor reconstruction of bone and misidentification of traumatic fractures as non-traumatic features in burned bones from forensic cases. Additionally, fragmentation (number and size) was inspected in this study but in real situations, fragmentation of burned bones is difficult to accurately determine and could result even from post-fire handling. Some fragments might be missing or lost in the fire debris, thus fragmentation number and size are not reliable indicators for HIF and BFT-fracture distinction – fragmentation pattern would be a better observation.
Albeit using defleshed bones for more uniformity of fracture creation and burning, the effect of soft tissue on the fracture features needs to be investigated too. Soft tissue retraction and muscle contraction can influence the type of HIFs formed and mimic/obscure any pre-burning trauma [1,30,31]. The checklist of features from this study makes the distinction of HIFs from BFT-fractures more efficient than relying on qualitative macro/micro features (colour, surface, outline and basic type of angle) or only metric features (dimensions, exact angle), but in a relatively controlled
29 setting. These results thus, need to be further investigated. Correspondingly, blind studies need to be performed [43] where all burned bones from this study would be inspected by 2 (or more) experts, to see if they can differentiate bones subjected to BFT-burning from burning-only. Then, similar blind studies need to be extended to other scenarios: for instance, bones from current study and analysis by less-experienced individuals (using characteristics features from this study as a guideline); macerated bones; fleshed bones (same burning duration and longer duration) and fleshed bones modelled in realistic forensic situations. Consequently, the features (and checklist) from present experimental research can be accurately evaluated for their usability and efficacy and eventual implementation in casework. Though the experiments were largely done in a controlled manner, the burning experiment was also designed to be more relevant to actual cases. The fire dynamics was not restrained by closing the windows or doors, or removing furniture already present in the firehouse – any influence on fire was caused by the air ventilation and surrounding environment [1,32,33].
The checklist utilized was constructed from combining commonly investigated features from prior studies, scrutinizing their corresponding definitions/descriptions followed by making these descriptions more detailed and specific. As such, less confusion occurs during analysis and more value is thus added to the usage of these features. Additionally, some features such as burning phases, fracture angle and dimensions were evaluated with different approaches as compared to former studies [3,7,9,11,16]. Both carbonization and calcination phases were sub-categorized as early and late to observe the burning pattern and other heat-induced changes of the bones with greater detail. Fracture angle was measured from photographs and a downward-slope across the fracture outline. This was to ensure a more consistent and systematic manner of angle measurements, preventing imprecise measurements from using a protractor on the bones, which tend to curve slightly on one side. The bone dimensions BFT and post-burning were measured in terms of length of fracture and ratio of length from tension to compression sides to confirm the fracture outlines (for example, transverse outlines showed ratios closer to 1), determine the shrinkage more easily and effect of fire on the fracture edges.
The setup for the burning experiment was a revision of those employed by Carroll & Smith (2018) [33], DeHaan, Campbell & Nurbakhsh (1999) [34] and Alunni, Grevin, Buchet & Quatrehomme (2014) [35]. The temperature reached by the wooden pyre (estimated from fuel specifications, 700C) associates with general house fires [1,36]. Calcination begins at this temperature and the results from this study follow this. Many environmental and individual (victim) factors affect the fire progression and resulting fracture features such as air ventilation, material of furniture/walls, climate for former and thickness of protective skin/soft tissue, anatomical area being burned, clothing for latter. These factors account for the varied exposure to fire, heat fluctuations over time and the consequently differing levels of colouration and burning in forensic fires/cremations [1,16,33]. As such, it is tough for experimental studies to entirely replicate the diverse forensic fire scenarios. The current study managed to mimic some casework-relevant conditions – bones surrounded by skin and clothing, specimens placed within a room of typical living room furniture and ventilation through doors and windows. Due to ethical reasons, pig skin was used instead of human skin and pig skin has mechanical properties similar to those of human skin hence, pig skin is a common substitute for human skin in experiments [36]. Practically, pig skin was also easier to obtain at the moment of experiment. Moreover, the skin to be used must not be decomposed or autolytic hence cannot be stored before use. As such, pig skin is more readily available and usable. In fires, radius-ulna bones are contracted upwards (pugilistic posture) allowing air and fire to surround the bones (all sides) [2]. Similarly, in this burning experiment, the forearms were surrounded by fire and air – this adds to forensic relevance of this study. The fire was extinguished using water spray, a common practice in forensic cases. However, it is important to note that the sudden thermal shock from the cold water to the bones will itself cause
