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This is to certify that the
thesis entitled

A MICROSPECTROPHOTOMETRIC CHARACTERIZATION
OF DYED AND BLEACHED HAIR

presented by

TIFFANY EDWINA CULMER

has been accepted towards fulﬁllment
of the requirements for the

MASTER OF degree in
SCIENCE

CRIMINAL JUSTICE

 

 

 

/ U Major i7rol‘essor’s Signature
April 4, 2005

 

Date

MSU is an Afﬁrmative Action/Equal Opportunity Institution

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WW

A MICROSPECTROPHOTOMETRIC CHARACTERIZATION OF DYED AND
BLEACHED HAIR
By

Tiffany Edwina Culmer

A THESIS
Submitted to
Michigan State University
in partial fulﬁllment of the requirements
for the degree of
MASTER OF SCIENCE

Department of Criminal Justice

2005

ABSTRACT

A MICROSPECTROPHOTOMETRIC CHARACTERIZATION OF DYED AND
BLEACHED HAIR

By

Tiffany Edwina Culmer

Microspectrophotometry was used to characterize dyed and bleached human hair
ﬁbers. The identiﬁcation of dyed hair is important when investigating a crime scene. If
bleaching or dyeing has artiﬁcially altered hairs, their evidential signiﬁcance will
increase. Microscopic techniques allow the analyst to examine and compare hair’s
structural characteristics but are rather limited for similar color comparison. Color is an
important feature in hair dye analysis because it helps in reaching conclusions about
similarities and differences.

This research proposed to determine how much dye is required for the visible
spectrum of one strand of hair to become consistent with that dye and to compare the
microspectra of bleached hair ﬁbers. The results of the hair dyeing analysis determined
that the even distribution of the dye varied with each brand name. The results of the hair
bleaching analysis demonstrated that there are no signiﬁcant spectral differences in hair

bleaching formulas.

ACKNOWLEDGEMENTS

“I can do all things through Christ who strengthens me...” Philippians 4:13. Nothing is
impossible when we put our trust in GOD!

To my parents, Edwin and Veronica Culmer, thank you for all the sacriﬁces that you
have made for me so that I can make my dreams a reality. You will always be the wind
beneath my winds.

To my boyfriend, Renard, thank you so much for the moral support you gave me every
step along the way.

To some very special ﬁiends, Bianca, Nadine and Sherri, thank you for always being
there. You made me laughed to help me keep my sanity and you listened.

To my advisor, Dr. Jay Siege], thank you for your advice and most importantly your
patience.

To, Dr. Paul Martin, thank you so much for providing this project for me.

iii

TABLE OF CONTENTS

LIST OF TABLES ................................................................................... vi
LIST OF FIGURES ................................................................................ vii
CHAPTER ONE: INTRODUCTION ............................................................... 1
The Structure of Hair ........................................................................ 2

Hair as Trace Evidence ..................................................................... 4

Hair Dyes ..................................................................................... 5

Hair Bleaches ................................................................................. 8

What is Melanin? ........................................................................... 10
Review of the Literature .................................................................. 11
Purpose of Study ........................................................................... 12
Limitations .................................................................................. 13
CHAPTER TWO: INSTRUMENTATION ...................................................... 13
History of Microspectrophotometry ..................................................... 13

Uses of Microspectrophotometry ......................................................... 13
Microspectrophotometry .................................................................. 1 5
CHAPTER THREE: METHODS AND MATERIALS ....................................... 18
Sample Preparation ........................................................................ 18
Preparation of Smears ..................................................................... 18

Hair Dyeing Process ....................................................................... 18

Hair Bleaching Process .................................................................... 21
Microspectrophotometric Analysis ...................................................... 22
CHAPTER FOUR: RESULTS AND DISCUSSION ........................................... 25
Results ....................................................................................... 25

Hair Dyed Samples ............................................................... 25

Hair Bleached Samples ........................................................... 28

Discussion ................................................................................... 29
CHAPTER FIVE: CONCLUSION AND FUTURE RESEARCH ........................... 32
APPENDICES ...................................................................................... 36
APPENDIX 1: SPECTRA OF DYE SMEARS ................................................ 37
APPENDD( 11: SPECTRA OF DYED HAIR FIBERS ....................................... 46
APPENDD( III: SPECTRA OF BLEACHED HAIR FIBERS .............................. 57

APPENDIX IV: PHOTOGRAPH LIBRARY ................................................. 59

APPENDIX V: LIST OF INGREDIENTS ..................................................... 71
NOTES .............................................................................................. 75
BIBLIOGRAPHY ................................................................................. 77

Table 1.

Table 2.

Table 3.

Table 4.

Table 5.

Table 6.

LIST OF TABLES

List of Brand Names and Colors of L’Oreal Dyes ............................ 20
Dye Processing Data .............................................................. 21
Bleach Processing Data ........................................................... 22
Results of Dye Amounts for Consistency ...................................... 27
Dye Color Differences Before and Aﬁer the Addition of Developer ...... 28
Racial Characteristics of Hair ................................................... 33

vi

Figure 1.
Figure 2.
Figure 3.
Figure 4.
Figure 5.
Figure 6.

Figure 7.

Figure 8.

Figure 9.

Figure 10.
Figure 11.

Figure 12.

Figure 13.

Figure 14.

Figure 15.

Figure 16.
Figure 17.
Figure 18.

Figure 19.

LIST OF FIGURES

Cross-Section of Human Hair ..................................................... 4
Stages of Permanent Hair Coloring .............................................. 8
Breakage of Double Bonds by Oxidizing Bleaches ........................... 9
Redox Reaction of Melanin ...................................................... 11
Basic Components of a Microscopical Spectrophotometer System ........ 15
0.5 ml Smear of L’Oreal Excellence Red Penny ............................. 38
0.5 ml Smear of L’Oreal F eria All-Out Red Copper

Red....... ................................................................................................ 39
0.5 ml Smear of L’Oreal F eria Double Intensity Auburn Red ............... 40
0.5 ml Smear of L’Oreal F eria Glowing Red-Hot Red ....................... 41
0.5 ml Smear of L’Oreal Preference Red Penny .............................. 42
0.5 ml Smear of L’Oreal Preference Smoldering Red ........................ 43
0.5 ml Smear of L’Oreal Preference Mega Reds Dark Intensity Copper
Red .................................................................................. 44
0.5 ml Smear of L’Oreal Preference Mega Reds Medium Intensity Red
Copper .............................................................................. 45
Representative Spectrum of Hair before Cosmetic Treatments

(Hair Fibers Used in Batches 1-14) ............................................. 47
Representative Spectrum of Hair aﬁer Bleaching

(Hair Fibers Used in Batches 1-8) .............................................. 48
Representative Spectrum of One Hair Fiber from Batch 1 @ 1.5 ml ...... 49
Representative Spectrum of One Hair Fiber from Batch 2 @ 1.0 ml ...... 50
Representative Spectrum of One Hair Fiber from Batch 3 @ 1.5 ml ...... 51
Representative Spectrum of One Hair Fiber from Batch 4 @ 1.0 ml ...... 52

vii

Figure 20.
Figure 21.
Figure 22.
Figure 23.

Figure 24.

Figure 25.

Figure 26.

Figure 27.
Figure 28.
Figure 29.
Figure 30.
Figure 31.
Figure 32.
Figure 33.
Figure 34.
Figure 35.
Figure 36.
Figure 37
Figure 38.
Figure 39.
Figure 40.

Figure 41.

Representative Spectrum of One Hair Fiber from Batch 5 @ 1.5 ml. . ...53
Representative Spectrum of One Hair Fiber from Batch 6 @ 1.0 ml. ....54
Representative Spectrum of One Hair Fiber from Batch 7 @ 1.5 ml. ....55

Representative Spectrum of One Hair Fiber from Batch 8 @ 1.5 ml. ....56

Overlay of Hair Fibers from Batches 9 —14 ................................... 58
Photo of Hair Fiber before Cosmetic Treatments

(Hair Fibers Used in Batches 1-14) ............................................ 60

Photo of Hair Fiber aﬁer Bleaching

(Hair Fibers Used in Batches 1-8) ....................................................... 60
Photo of Hair Fiber from Batch 1 @ 1.0 ml of dye .......................... 61

Photo of Hair Fiber from Batch 1 @ 1.5 ml of dye .......................... 61

Photo of Hair Fiber ﬁ'om Batch 2 @ 1.0 ml of dye .......................... 62
Photo of Hair Fiber from Batch 3 @ 1.0 ml of dye .......................... 62

Photo of Hair Fiber from Batch 3 @ 1.5 ml of dye .......................... 63

Photo of Hair Fiber from Batch 4 @ 1.0 ml of dye .......................... 63

Photo of Hair Fiber from Batch 5 @ 1.0 ml of dye .......................... 64
Photo of Hair Fiber from Batch 5 @ 1.5 ml of dye .......................... 64

Photo of Hair Fiber from Batch 6 @ 1.0 ml of dye .......................... 65

Photo of Hair Fiber from Batch 7 @ 1.0 ml of dye .......................... 65

Photo of Hair Fiber from Batch 7 @ 1.5 ml of dye .......................... 66

Photo of Hair Fiber from Batch 8 @ 1.0 ml of dye .......................... 66

Photo of Hair Fiber from Batch 8 @ 1.5 ml of dye .......................... 67

Photo of Hair Fiber from Batch 9 ............................................... 67
Photo of Hair Fiber from Batch 10 ............................................. 68

viii

Figure 42.
Figure 43.
Figure 44.

Figure 45.

Photo of Hair Fiber from Batch 11 ............................................. 68

Photo of Hair Fiber from Batch 12 ............................................. 69
Photo of Hair Fiber from Batch 13 ............................................. 69
Photo of Hair Fiber from Batch 14 ............................................. 7O

CHAPTER ONE: INTRODUCTION

Hair evidence is important because it is commonly left at a crime scene without
the subject’s knowledge. Hair ﬁbers have unique shedding and transfer properties. Hair
ﬁbers are constantly shedding and are not easily lost or displaced from fabric and
clothing. The types of hair recovered, their condition and number of hairs found all
impact their value as evidence in a criminal investigation. The victim’s and/or suspect’s
cosmetically treated hair will inﬂuence the signiﬁcance of the case. Color is the most
comparative characteristic available to a forensic examiner.

Hair association made between a suspect and a victim or a suspect and a crime
scene can potentially provide very valuable and persuasive evidence in a courtroom. Hair
can provide crime investigators with important clues. Apart from burning, hair is virtually
indestructible. It remains identiﬁable even on bodies in an advanced state of
decomposition or attached to objects after a crime has been committed.

Hair evidence may be subjected to microscopic examination to determine
physical characteristics, biological examination to determine DNA type and toxicological
examination to determine the presence of drugs or poisons.

Microscopic analysis is straightforward. Several types of microscopes can be used
including stereoscope, compound and comparison.

Stereoscopic microscopy provides a broad overview of the range of characteristics
of hairs. They have low magniﬁcation capabilities. The compound or polarizing
microscopes have a higher magniﬁcation. As a result they provide more details about the

hair’s structural characteristics. Ifthere is a need to make a side by side comparison

between a questioned and known hair sample, a transmitted light comparison microscope
is used.

In 1985, Robertson and Aitken conducted a study on the value of microscopic
features in the examination of human hairs.l The study concluded that microscopic
techniques allow the analyst to compare hair’s structural characteristics but are rather
limited for color comparison.

Visible microspectrophotometry is used to determine the exact color of a material.
It is especially useful when comparing two objects that are similar in color. The color of a
dyed hair is its most important characteristic. The human eye has the capability to
discriminate color better than a microspectrophotometer however, there are a number of
problems in the visual comparison of colors under a microscope. It is subjective because
of the differences in color sensitivity of the observer and also because of the variations in
viewing conditions within the microscope. These variations are due to slight differences
in mirrors, optical glass colors and polarizers.2

The identiﬁcation of dyed hair is important when investigating a crime scene. If
bleaching or dyeing has artiﬁcially altered hairs, their evidential signiﬁcance will

increase. In this study, human head hairs were analyzed by microspectrophotometry in an

attempt to characterize dyed and bleached hair.

The Structure of Human Hair

Each strand of hair grows out of a tiny pocket in the skin called a follicle. Hair is
composed of dead cells that are ﬁlled with insoluble proteins and held together by strong

attachments between the cells. These proteins are hard ﬁbers called keratin. Each hair

has a root and shaft within its follicle. The growth of human hair is cyclic, which involves
three stages. Anagen is the stage of active growth. The root is attached to the hair follicle
for continued growth. This stage can vary from two to six years. The ﬁnal resting stage of
the hair growth cycle is the telogen stage. This stage normally lasts ﬁve to six weeks. In
this stage, the hair is pushed out of the follicle and naturally sheds. The transitional
period between the growth and the resting stages is known as the catagen stage. This
transitional stage can last for one to two weeks. In this stage, hair growth continues but at
a very slow rate. The size of the hair root bulb begins to shrink as it is being pushed out
of the hair follicle. There is a stop of protein synthesis as the hair follicle retreats towards
the surface.

Hair is continually shed and renewed. Growth is not synchronized and each hair
passes through the three phases independently. The average human scalp has 100,000
hairs and an average of 30 to 100 hairs are lost per day by shedding.

Hair has three layers: cuticle, cortex and medulla. The cuticle is the outermost
layer of the hair strand. It serves as a protective sheath around the hair, which is
comprised of overlapping scales. It is often compared to roof tiles.

The cortex is made up of spindle-shaped cortical cells, known as keratin, that are
aligned in a regular array, parallel to the length of the hair. Single keratin molecules
consist of a combination of amino acids, in particular cystine, which form stable
disulphide bridges between molecular chains. These disulphide bonds form down the
length of the keratin chains, joining them together like rungs of a ladder. Pigment
granules containing melanin are embedded within the cortex. These melanin-ﬁlled

granules are scattered throughout the cortex of the hair. There is no set pattern and no set

amount. This is how nature creates so many variations of hair color. The distribution,
shape and color of these granules provide points of comparison.

The medulla is a shaft that runs through the middle of the hair. The medulla can
either be continuous, interrupted, fragmented or absent. One important aspect of the
medulla is that the presence and appearance vary from individual to individual and even
between hairs of a given individual. It may also differ from one area of a hair strand to

another.

Figure l — Cross-Section of Human Hair3

   
  
  

M e dulla
C ortex

Cuticle

Scales

Hair as Trace Evidence

Edmund Locard ﬁrst recognized the value of ‘trace’ forensic evidence in 1910
when he proposed his 'exchange theory'. It simply stated that 'every contact leaves a
trace'. It means that anybody entering a crime scene will take something of that crime

scene with them, as well as leaving something behind. This can constitute something as

obvious as a ﬁngerprint or a shoe impression. It can even be microscopic evidence such
as hairs or a piece of ﬁber from clothing.

Shed human hairs are one of the most commonly secured biological evidence
materials at crime scenes.4 Hair often can be found on the ﬂoor near the weapon or point
of impact between suspect and victim. These hairs may have fallen out naturally or
forcibly removed, which may suggest a violent confrontation. Human hair is useful
associative evidence because it originates directly from the individual.

The analysis of hair typically involves the inspection by microscopic techniques
to determine distinguishing characteristics that may help to associate an individual to the
hair sample collected. However, absolute personal identiﬁcation cannot be provided from
microscopic analysis of hair evidence. If hair is pulled out during the anagen phase, the
root may be surrounded by a translucent tissue that is known as the follicular tag. If
nuclear DNA can be extracted, it can lead to the identiﬁcation of an individual. In
addition, recent advances in mitochondrial DNA (mtDNA) has enabled genetic
characterizations of small hair sample with or without root cells.5

Forensic analysts are often asked to compare hair found at a crime scene with hair
from a particular individual. Some factors that must be taken into account by the
examiner include; species, racial origin, body location, the presence of cosmetic

treatments and the manner in which the hair was removed.

Hair Exes
Chemistry is the basis behind all cosmetics. It is through modern chemistry that

has made it possible to change hair color for a day or forever. Hair dyes can be compared

to paint because they cover hair strands by chemical coloring or by mixing with the
melanin granules without altering them. The main components in most hair dyes are
phenylenediamines, aminophenols, dihydroxybenzenes, nitro compounds and their salts.
These dyes facilitate simple and rapid hair coloring.6 There are three general categories of
dyes on the market. They are deﬁned by the duration the color remains on the hair.

Temporary dyes (rinses) comprise of water-soluble acid dyes and water-soluble
pigments, which are deposited on the surface of the hair with no penetration into the
cortex. These dyes do not lighten or change the structure of the hair. The color is
removed within one to three shampoos.

Semi-permanent dyes contain medium sized, basic molecules. These molecules
contain simple derivatives of nitroanilines, nitrophenylenediamines and
nitroaminophenols. The pH of the dye causes a minor alkaline reaction to take place that
swells the hair shaft and causes the cuticle to rise. This allows some of the dye molecules
to enter and diffuse throughout the cortex. These dyes cause mild chemical and physical
changes in the hair shaft. They usually wash out after ﬁve to ten shampoos. Semi-
permanent dyes come in the form of liquid, gel or aerosol foam.

Permanent dyes are marketed as two component kits. One component is a
combination of dye precursors or intermediates (such as 2,5-diarninotoluene, N,N-bis(2-
hydroxymethyl)-p-phenylenediamine and p-aminophenol) and couplers (such as
resorcinol, chlororesorcinol, methyl resorcinol and m-aminophenol) in an alkaline base.
The other component is a stabilized solution of hydrogen peroxide. The two components
are mixed together prior to use. The precursors and peroxide diffuses into the hair shalt

by an alkaline reaction that causes the cuticle to swell. The hydrogen peroxide oxidizes

the melanin and lightens the color of the hair. This lightening action depends on three
things: the strength of the developer, the level of the ammonia in the product and the
processing time.

Peroxide breaks chemical bonds in the hair, releasing sulphur, which accounts for
the characteristic odor of hair color. As the melanin is decolorized, a new permanent
color is bonded to the cortex.

The cuticle closes and the color molecules are trapped inside with the help of a
neutral or slightly basic shampoo which stops the alkaline reaction. These dyes damage
the hair shaft by increasing its porosity causing it to become harder to untangle.
Permanent dyes cannot be washed out with shampoo, however the color can fade with
repeated shampooing.

Hair should be bleached before dyeing if the color of the hair is darker than the

desired hair color.

Figure 2 — Stages of Permanent Hair Coloring7

 

 

Colorless molecules Forming large . _
plus developer water-insoluble Which remain .
penetrates hairshaﬁ. .. colored pigments. . .. trapped 1n the hair.

Hair Bleaches

Hair bleaching is a process of lightening the hair by changing part or all of the
melanin pigment in the cortex into a colorless substance. This process occurs in two
steps: the degradation of the granules and the decoloration of the solubilized pigments.8
In most cases, bleaching is used to prepare the hair for dyeing. During bleaching the
melanin pigment undergoes irreversible physiochemical changes which result either in

the lightening or complete elimination of the original ﬁber color.9

All natural hair colors are created from two types of melanin: eumelanin (black
pigment) and pheomelanin (red/yellow pigment). The type of melanin and the size of the
granules determine whether hair will be black, brown, blonde or red. The amount of
melanin and its distribution determine how dark or light the hair color will be.

Black hair is created from granules full of eumelanin densely packed in the hair's
cortex. Brown hair, depending on its darkness or lightness, is created from granules ﬁlled
with eumelanin and more sparsely distributed along the cortex than those of black hair.
The red/yellow pheomelanin is believed to cause the warm, golden, or auburn tones
found in most brown hair. Granules ﬁlled with pheomelanin create red hair. The
pheomelanin in red hair is less densely packed in its granules. Its shape is somewhat more
irregular than eumelanin. It is slightly rounder and more diﬁ‘used. Gray hair is caused by
the gradual reduction of melanin production over time. The body produces less and less
melanin, and the result is a loss of color strength. Some hairs may reach a total absence of
color (white).

Hydrogen peroxide is one of the most common lightening agents. It is a very
powerful oxidizing agent because it reacts directly with double bonds in large organic
molecules to form organic peroxides. Double bonded molecules have a tendency to
absorb light and therefore provide the molecule with its color. When the double bonds are

destroyed, the color is removed.

Figure 3 - Breakage of double bonds by oxidizing bleacheslo

H O O O 0
II I [or II II II II
H—C—C=C—C—H —> H—C—C—OH + OH—C—C—H

H O Colorless

All bleaching methods are oxidative-alkaline treatments. 11 The solution that is
used to bleach hair contains alkali to soften the cuticle and hydrogen peroxide to release
oxygen. This oxygen penetrates the hair and reacts with the natural pigments in the
cortex, oxidizing them and breaking them down. The melanin is still present, but the
oxidized molecule is colorless. Bleached hair tends to have a pale yellow tint however,
the degree of color left after bleaching depends on the production of eumelanin and
pheomelanin pigments. The yellow color is the natural color of keratin, the structural
protein in hair. The bleach mixture also contains modiﬁers to slow the release of oxygen

so that the reaction happens over a period of about an hour.

What is Melanin?

Melanin is the natural hair color pigment in the cortex layer. Specialized cells,
called melanocytes that are contained in the papilla of the follicle produce melanin
granules. Melanin is a family of polymers that are formed as an end product during the
metabolism of the amino acid tyrosine. As a result of its high molecular weight, this color
cannot be changed except by intense oxidation or concentrated alkaline solutions.

Melanin is an ill deﬁned and an insoluble macromolecule with many alternating
single and double bonds. This organic molecule has the ability to absorb light at many
frequencies. Melanin absorbs light energy very efficiently and disperses it as heat.

Color in organic materials is the result of light absorption by certain chemical
conﬁgurations called chromophores in molecules. C = C and C = 0 bonds are examples
of chromophores. A chromophore is a part of a molecule that is able to absorb UV or

visible light thus producing color in organic compounds.

10

Figure 4 - Redox Reaction of Melanin12

Melanin +

H202

 

"O

~CH-C—NH-i

R

~+H202 —'1

 

”‘0

FCH-C-NH-

P

 

 

 

 

 

 

 

Wheren=m+p

Review of Literature

Degraded Melanin

7

II
—CH-C-NH—

 

 

 

 

There are many published journal articles on cosmetically treated hairs. Past

research have utilized methods such as stereoscopic microscopy, polarized light

microscopy (PLM), ultraviolet light microscopy, scanning electron microscopy

(SEM/EDX), Fourier transform infrared spectroscopy (F T-IR), gas chromatography-mass

spectrometry (GC-MS), pyrolysis gas chromatography (PGC), thin layer chromatography

(TLC), microspectrophotometry and secondary ion mass spectrometry (SIMS) to

determine physical and chemical properties. This research was necessary to provide a

qualitative and quantitative comparison of permanent hair dyes and hair bleaches.

N. Tanada et al. have done extensive research of oxidation dyes on human hair.

In one study, N. Tanada et a1. purposed to improve the reliability of identiﬁcation of dyed

hair in addition to conventional procedures, using gas chromatographic-mass

spectrometric analysis (GC-MS) and secondary ion mass spectrometry (SIMS) to analyze

nine common components of permanent hair dyes. It was concluded that hair dye

ll

component analysis of a dyed hair specimen could serve as an index for use in forensic
hair comparison.

Another study conducted by N. Tanada et al. involving selective ion monitoring
(SIM) analysis demonstrated that stained hair conﬁrmed to be very beneﬁcial for the
identity of human hair in practical cases. The ﬁve identiﬁable components in this study
were, p-phenylenediamine, toluene-2,5-diamine, o-arninophenol, m-aminophenol and p-
aminophenol. The peak area percentage of the aminophenols versus diarnines is thought
to be effective for the identiﬁcation of the brand of oxidation hair dyes.”

A comprehensive study on the mechanism of hair bleaching was conducted by
Wolﬁam et al. This investigation aimed at obtaining a better understanding of the
complex processes that are associated with both the melanin pigments and hair proteins.
This study employed both microspectrophotometry and thin-layer chromatography
techniques. Microspectrophotometry was utilized to obtain the visible and UV spectra of
melanin and thin-layer chromatography was employed to separate the melanin oxidation
products. It was discovered in this study that the solubilization of melanin by hydrogen
peroxide is the ﬁrst step in the reaction sequence. It also showed that prolonged
treatment resulted in the decolorization of hair ﬁbers. The reaction between keratin and

hydrogen peroxide is conﬁned mainly to the cystine residues.

Purpose of Study

This study is an attempt to determine the utility of microspectrophotometry as an
analytical tool in the characterizing of permanently dyed and bleached hair. There are two

purposes of this study. The ﬁrst is to determine if the amount of dye that is required for

12

one strand of hair to become consistent varies between dyes that are all produced by the
same manufacturer, L’Oreal.

The second purpose is to determine whether bleaching is spectrally unique. In
this study, the SEE 1100 was used to obtain the absorbance spectra in the visible regions

of hairs that were treated with permanent dyes and bleaches.

Limitations
This study focuses exclusively on the analysis of Caucasian hair. This is clearly
an artiﬁcial concept in casework situations as the population of most countries is of

mixed ethnic origin.

CHAPTER TWO: INSTRUMENTATION

History of Microspectrophotometry
Caspersson reported the ﬁrst literature reference, which described the

combination of a microscope to a spectrophotometer between 1925-1940. The
spectrophotometer consisted of a sector photometer, monochromator, and a photocell. A
microscope with quartz optics permitted both transmission of the source (visible,
ultraviolet, or ﬂuorescence) and visual examination of the sample.

With the emergence of lens technology, the Cassegrainian objective provided a
high quality visual image, reasonable magniﬁcation of up to 18x, and transparence in the
UV, visible, near and mid-IR regions. The instrumental advances in the development of
ultraviolet microscopy during the 19405 laid the foundation for commercial UV
microscopy. A quality spectrum could be obtained within several minutes to several

hours depending on the size and sensitivity of the specimen.

Uses of Microspectrophotometg

This instrument was primarily introduced for the identiﬁcation of tissue section
and single cells in biological studies. Today this analytical technique is being utilized
extensively in many areas. Quantitative absorbance data are attainable for many
applications in the ﬁeld of forensic science, speciﬁcally in the area of trace evidence. A
forensic examiner can employ this technique to analyze ﬁbers, paint chips and questioned
documents. This technique is also being applied to the cosmetic industry to help provide
an understanding of how cosmetic treatments affect human hair properties. This

knowledge is critical for the development of successful products.

14

Microspectrophotometry

A microspectrophotometer is an instrument that is a combination of a microscope
and a computerized spectrophotometer. A spectrophotometer measures the intensity of a
light beam before and after it passes through a sample and compares these two intensities.
Spectrophotometry is based on the wave theory of light and the principle that substances
selectively absorb certain wavelengths and reﬂect or transmit others. It provides an
absolute measurement because it compares the light that has been transmitted or reﬂected
at each wavelength with the intensity of light incident on the object, making the result

independent of the type of illumination used.14

5

Figure 5 - Basic Components of a Microscopical Spectrophotometer System.l

 
   

 

  
 
 

Dispersive
system

   

Microscope Detector

 

 

 

A pictorial representation of the manner in which a certain colored object reacts to
various wavelengths of light is referred to as a spectrum. Microspectrophotometry has
been widely utilized in forensic examination of trace evidence for objective evaluation of
colors of minute materials. ‘6 Different substances react to light in different ways. The
spectra of different objects can be compared to each other to determine similarities and

differences.

15

An advantage of using microspectrophotometry as opposed to comparing colors
by the eye via microscopy is that phenomenon of metarneric colors is eliminated.
Metamerism occurs when two colors match in one light but fail to match in another. The
spectral comparisons that are produced using microspectrophotometry provide more
improved and objective conclusions than by the comparison made by visual examination.

In this study, the SEE 1100 microspectrophotometer was utilized in conjunction
with the Grams/32 program software. Grams/32 is an integrated suite of instrument
controls, spectral data analyses programs and a fully relational database. To compare the
precise color of the hair dyes in this project, the transmittance method was employed
utilizing a halogen lamp.

Combined with its high discriminating power, its non-destructive and rapid
method of analysis that requires little sample preparation, microspectrophotometry can
provide excellent results. Data ﬁom both instruments are stored as spectra. The
instruments can load up to thirty spectra in the memory stack. These stored spectra can
then be retrieved for future comparison if it is necessary.

The determination and accuracy of the instruments were determined by routine
calibrations with the aid of ﬁlters. The wavelength accuracy is determined through
holrnium oxide and didymium ﬁlters. The spectra of these ﬁlters display sharp peaks at
well-documented wavelengths. The holrnium oxide ﬁlter is used to calibrate wavelength
from 280 nm to 640 nm in the visible and UV regions. The didymium ﬁlter has peaks in
the visible and near IR wavelength regions, which is used to calibrate wavelengths from
440 nm to 880 nm. The photometric accuracy is determined through a sequence of

neutral density ﬁlters (OD = 0.1, 0.5, 1.0). These ﬁlters are characterized by a ﬂat optical

l6

response in the wavelength region from 250 nm to 1000 nm. Each ﬁlter comes with NIST
traceable wavelength values. The NIST traceable set that is provided by SEE
Incorporated adheres to the ASTM standard practices for describing and measuring the

performance of UV, visible and near IR spectrophotometers.

l7

CHAPTER THREE: METHODS AND MATERIALS
Sample Preparation

All of the hairs used in this study were head hairs cut from one Caucasian female
individual with clean scissors and stored, until use, at room temperature. The hair ﬁbers
had only been subjected to normal wear, which included shampoos, conditioners, water
settings, hair dryers, combing, brushing and atmospheric exposure. The hair ﬁbers were
not chemically treated prior to this experiment. The samples were washed twice with a
non-conditioning commercial shampoo to eliminate external contaminates and allowed to
dry at room temperature for 3 days. The hair samples were divided into two equal batches

and were kept separate from each other.

Preparation of Smears
Eight bottles of permanent red hair dyes were purchased from a local beauty

supply store. Table 1 lists the brands and colors of the dye. 0.5 ml of each dye was added
to its corresponding developer in a ratio of 1:1. The solutions were thoroughly mixed and
allowed to process. Small amounts of each mixture were smeared on glass microscope
slides and were allowed to dry at room temperature. Once dried, ﬁve sample scans were
run on each sample in different areas of the hair dye smear. Images in this thesis are

presented in color.

The Hair mung Process
In order to establish practical and reliable dyeing and bleaching methods, the

manufacturers’ instructions were carefully followed. Initially, the hairs were bleached

prior to dyeing because the present hair color was much darker than the desired hair

18

color. Hair ﬁbers were stained with eight permanent hair dyes. Once the hair ﬁbers were
bleached, they were evenly separated into batches. Hairs were randomly selected out of
each batch and nine sample scans were obtained from different locations on the hair. The
spectra from the scans were then overlaid and printed for further examination.

The hair bundle was composed of dark brown virgin hair 5 cm in length. The hair
ﬁbers were bleached in order to lighten the natural hair shade and to prepare it for dyeing.
14 g of a bleaching powder, L’Oreal Quick Blue, was added to 29 ml of its corresponding
developer. Table 2 lists the names of the bleaches and their corresponding developers.
The powder and developer were thoroughly mixed and applied to the hair using the foil
wrap technique. The hair was placed on a small piece of aluminum foil and covered with
the bleach mixture. The foil was then folded into a small package and processed to until
the hair was white in color. The hair was checked every 10 minutes to determine the ﬁnal
color. When the desired shade was obtained, the hair ﬁbers were carefully rinsed with
water, sharnpooed and air-dried. These hair samples were divided up into 8 different
batches that each contained 50 strands of hair and were assigned numbers 1 to 8.

Batches 1 to 8 were now ready to be dyed. The hair ﬁbers were stained with lml
of dye according to the manufacturer’s directions and were stored at room temperature
until the time of analysis. Five hair ﬁbers were randomly selected from each batch and
were individually mounted on using microscope slide/cover slip preparations in 1.539
Cargille refractive index ﬂuid.

Nine sample scans were obtained from different locations on each hair ﬁber. The
scans were overlaid and examined for consistency. If the color was not consistent the

remaining hair ﬁbers in all batches were stained with the dye corresponding to its batch

l9

number in increments of 0.5 ml. This process was repeated until the dye had been equally
distributed throughout the hair ﬁbers. The spectra were overlaid and printed for further

examination.

Table 1 - List of Brands and Colors of L’Oreal Dyes

 

 

 

 

 

 

 

 

 

 

 

 

Brands of L’Oreal Dyes Name of Color

Excellence Red Penny

Feria All-Out Red Copper Red
Féria Double Intensity Auburn Red
F éria Glowing Red-Hot Red
Preference Red Penny

Preference Smoldering Red

Preference Mega Reds Dark Intensity Copper Red
Preference Mega Reds Medium Intense Red Copper

 

20

Table 2 —- Dye Processing Data

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Batch L’Oreal Color Developer Processing
Number Brand Time (min)
1 Excellence Red Penny L’Oreal 20 Vol. 25

Oreor Creme
Developer Oxydant
2 Feria All-Out Red L’Oreal Féria Multi- 25
Copper Red Tone Color
Developer Plus
3 Féria Double Intensity L’Oreal Féria Multi- 25
Auburn Red Tone Color
Developer Plus
4 Féria Glowing Red- L’Oreal Féria Multi- 25
Hot Red Tone Color
Developer Plus
5 Preference Red Penny L’Oreal 20 Vol. 25
Oreor Creme
Developer Oxydant
6 Preference Smoldering Red L’Oreal 20 Vol. 25
Oreor Creme
Developer Oxydant
7 Preference Dark Intensity L’Oreal 20 Vol. 25
Mega Reds Copper Red Oreor Creme
Developer Oxydant
8 Preference Medium Intense L’Oreal 20 Vol. 25
Mega Reds Red Copper Oreor Creme
Developer Oxydant

 

The Hair Bleaching Processes

Six commercially available bleaching formulas for home use were individually

applied to the batches of hairs, labeled numbers 9 to 14, according to the manufacturer’s

instructions via the foil wrap technique. Refer to Table 3 for bleach processing data.

Five hair ﬁbers were randomly selected from each batch and analyzed by wet-mount

comparison with 1.539 Cargille refractive index ﬂuid mounting media. Five sample scans

were obtained ﬁ'om different locations on each hair. The scans were overlaid and

21

 

examined for similarities and differences. The spectra were overlaid and printed for

further examination.

Table 3 — Bleach Processing Data

 

 

 

 

 

 

 

 

 

 

 

 

 

Batch Powder Developer Amount of Amount of Processing
Number Bleach Powder Developer Time
Bleaﬂ) (ml)
9 Clairol Salon Care 20 8 17 20
Kaleidocolors Volume Creme
10 Clairol BW2 Clairol 30 8 13 50
Volume
Clairoxide
11 L’Oreal L’Oreal 30 8 17 50
Quick Blue Volume Oreor
Creme
Developer
Oxydant
12 L’Oreal L’Oreal 30 8 19 50
Super Oréal Volume Oreor
Blane Creme
Developer
Oxydant
l3 Salon Care Clairol 30 8 13 50
Quick White Volume
Clairoxide
14 Wella Wellite L’Oreal 30 8 16 50
Volume Oreor
Creme
Developer
Oxydant

 

Microspectrophotometric Anaﬂsis

II"

A L a
r r

‘ ic analysis was performed on the SEE 1100 (version 2.0)

in the forensic science laboratory at Michigan State University. The initial calibration of

the instrument was performed prior to the initiation of this microspectrophotometric

22

 

analysis. Each ﬁlter was measured 50 times over the course of 25 days in order to
establish a baseline average for each speciﬁc microspectrophotometer.

The instrument was calibrated before the dye smears and hair mounts were
analyzed. Prior to calibration, there was a stabilizing period for the instruments of 30
minutes. The ﬁrst component to be calibrated was the microscope. It was adjusted for
Koehler illumination. This term refers to a type of optically centered and focused
illumination that is essential to the transmission of the maximum amount of analytical
beam energy through the system.17

It was necessary that all the wavelength measurements stayed within the
designated range of i 3.0 nm of the National Institute of Science and Technology (N IST)
values on the calibration certiﬁcates when the same measurement conditions are utilized.
All photometric measurements were also required to remain within the range that is
speciﬁed on the certiﬁcates when the same measurement conditions are utilized. The
instruments were calibrated with the NIST Traceable Filter Set on a daily basis prior to
use of the instruments. The calibrations were archived for future instrument evaluation.

The instruments had to go through another series of steps before a sample
spectrum can be taken. The ﬁrst step was to apply the autogain function in order to
optimize the instruments. This feature automatically adjusts the instrument. The light
source shutter was closed and a dark scan was taken. This was done to ensure that the
microspectrophotometers were free of light leaks and were not affected by stray light.
The light path was opened and the reference scan was taken. This scan is performed
within the ﬁeld of area that does not contain the sample. The sample was then positioned

so that it could be analyzed. The sample scan button was pressed and the transmittance

23

(%T) spectra option was chosen. A reference scan is always obtained before every

sample scan that is taken outside of the previous reference area.

24

CHAPTER FOUR: RESULTS AND DISCUSSION

Results

Hair I_)yed Samples

Upon comparison of the spectrum of the dye smears, it was observed that the
spectrum varied slightly in shape and relative peak heights. The wavenumbers of the
most predominant peak in the microspectrophotometry spectrum for the 0.5 ml dye
smears were compared to each other. The spectra were all consistent with each other (see
Figures 6 — 13), with some variation. The range of the maximum absorbance peaks
ranged from 468.9 — 481.2. This is a difference of 12.3 absorbance units. Some of this
variation could have resulted from the manual markings of the peaks.

The amount of dye that was required to obtain equal distribution throughout one
strand of hair was determined by microspectrophotometry. There were little variations in
intensity across the spectral region as expected due to dye diffusion across the hair ﬁber.
This reinforces the importance of taking multiple scans of one object in order to
determine the overall variation among spectra.

The evaluation of the recorded spectra obtained by microspectrophotometry
determined that the amount of dye needed to obtain a consistent color on a hair ﬁber
varies with the product. The amount of dyes that was required for consistency are listed
in Table 3.

The nricrospectrophotometry absorbance spectra of the dyed hair ﬁbers were
compared and classed based upon the overall shape of the spectral curve. Feria All-Out

Red Copper Red, F eria Glowing Red-Hot Red, Preference Mega Reds Dark Intensity

25

Copper Red and Preference Mega Reds Medium Intensity Copper Red spectrum could
not be distinguished from each other.

Excellence Red Penny and Preference Red Penny spectrum also could not be
distinguished from each. Feria Double Intensity Auburn Red and Preference Smoldering
Red spectrum were the only two dyes that were easily identiﬁable from all the other dyes
analyzed in this study.

It was predicted that the products that had the same sub-brand should have
required the exact amount of dye to produce consistency, however this study did not
support that hypothesis in all cases. There were four sub-brands of dyes that were
manufactured by L’Oreal that were analyzed in this study. They were Excellence, Feria,

Preference and Preference Mega Reds.

26

Table 4 - Results of Dye Amounts for Consistency

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Batch L’Oreal Name of Developer Amt of Dye
Number Brand Color (ml) for
Consistency
1 Excellence Red Penny L’Oreal 30 Vol. 1.5
Oreor Creme
Developer Oxydant
2 F éria All-Out Red L’Oreal Feria Multi- 1
Copper Red Tone Color Developer
Plus
3 Féria Double L’Oreal Féria Multi- 1.5
Intensity Tone Color Developer
Auburn Red Plus S
4 Feria Glowing Red- L’Oreal Feria Multi- 1
Hot Red Tone Color Developer
Plus
5 Preference Red Penny L’Oreal 30 Vol. 1.5
Oreor Creme
Developer Oxydant
6 Preference Smoldering L’Oreal 30 Vol. 1
Red Oreor Creme
Developer Oxydant
7 Preference Dark Intensity L’Oreal 30 Vol. 1.5
Mega Reds Copper Red Oreor Creme
Developer Oxydant
8 Preference Medium L’Oreal 30 Vol. 1.5
Mega Reds Intense Red Oreor Creme
Copper Developer Oxydant

 

Although all of the dyes examined in this experiment were marketed as red in
color, there were color differences before and after the addition of the developer. These
color differences are listed in Table 4. This proves that that there are many different

formulas that are used to alter hair color.

27

Table 5 - Dye Color Differences Before and After the Addition of Developer

 

 

 

 

 

 

 

 

 

 

 

 

 

 

L’Oreal Name of Developer Color of Color of
Brand Color Dye Mixture
Excellence Red Penny L’Oreal 30 Vol. Light Dark Purple

Oreor Creme Yellow
Developer Oxydant
Féria All-Out Red L’Oreal Feria Light Dark Purple
Copper Red Multi-Tone Color Brown
Developer Plus
F éria Double L’Oreal Féria Light Dark Purple
Intensity Multi-Tone Color Brown
Auburn Red Developer Plus
Feria Glowing Red- L’Oreal F éria Light Dark Purple
Hot Red Multi-Tone Color Brown
Developer Plus
Preference Red Penny L’Oreal 30 Vol. Light Dark Purple
Oreor Creme Brown
Developer Oxydant
Preference Smoldering L’Oreal 30 Vol. Light Dark Purple
Red Oreor Creme Brown
Developer Oxydant
Preference Dark Intensity L’Oreal 30 Vol. Light Dark Purple
Mega Reds Copper Red Oreor Creme Brown
Developer Oxydant
Preference Medium L’Oreal 30 Vol. Light Dark Purple
Mega Reds Intense Red Oreor Creme Brown
Copper Developer Oxydant

 

Hair Bleached Samples

This study was based more on quality rather than quantity. The hair ﬁbers were

separated into batches 9-14 however, the number of hair ﬁbers in each batch were not

always equal as those in the hair dyeing process. Each batch was bleached with a

different bleaching formula using the foil wrap technique. Hairs were randomly selected

out of each batch and ﬁve sample scans were taken from different locations on each hair.

28

 

After representative hair samples were scanned from each batch, the most compatible
spectra was chosen and overlaid.

Figure 24 shows an overlay of hair samples from batches 9-14. All six of the
bleaching formulas demonstrated a similar spectral pattern. The evaluation of the
transmission spectra obtained from analyzing the bleached hair ﬁber samples was based
upon the peak maxirna and overall shape of the spectral curve. All the bleaching formulas
used in this spectral comparison showed that there is no spectral uniqueness about any

particular bleaching formula.

Discussion

Forensic analysis of hair centers on color and structure that is determined through
microscopic magniﬁcation. Head hairs are usually the longest hairs on the human body.
These hairs are subjected to more alterations than hairs from any other body part. The
most popular hair treatments are bleaching and permanent dyeing. Nature develops its
own precursors within melanocytes, as a result the effects of hair treatment depend on the
characteristics of the individual hair ﬁbers.

The primary purpose of this study was to determine if the amount of dye that is
required for one strand of hair to be consistent varies between permanent dyes that are
produced by the same manufacturer. The quantitatively determined values were achieved
by adding increments of 0.5 ml for the amount dye as needed.

When it comes to normal hair care coloring, permanent color is the most popular
choice because it lasts the longest and gives the most dramatic change of tone. Hair care
products are available in permanent hair colors that will last until the hair is cut and/or the

roots grow out.

29

Permanent hair dyes start out as colorless molecules that are small enough to
squeeze between the cuticle cells and get inside the hairs. These tiny molecules react with
hydrogen peroxide to become colored molecules. They also form clusters that are too
large to wash out of the hair, leaving your hair permanently colored. A hair color can’t be
judge by looking at the dye mixture. The color of the pigments will change as the
colorless molecules react with hydrogen peroxide. At the end of a successful coloring
operation, the giant pigment molecules are securely in place behind the translucent
cuticle, which is once again protecting the cortex.

Permanent hair dyes are also referred to as oxidative hair dyes. Hydrogen
peroxide in the oxidative hair dye formulae serves as a bleaching agent for the natural
pigment of the hair. The color formation is dependent on precursors present in the dyeing
solution, its pH and the time of contact with the hair. ”"9 Oxidative hair dyes are resistant
to fading by sharnpooing, but re-coloring of hair is required approximately every six
weeks due to hair growth. The time of contact of the dyeing solution on the hair can vary
from 15 - 50 minutes. Permanent hair dyes were chosen because these products provide a
lasting color effect, 100% grey coverage and represent a large percentage of the hair
coloring market.

A comparison of all the active ingredients of each dye used in this study was
done. The result corresponded with the ﬁndings from the analysis that although these
dyes were marketed as the same color, they were different in their chemical composition.
The components in all of the dyes (see Appendix V) were listed on the dyes’ labels,
however there were a few active ingredients that were either present and/or absent in the

some of the dyes. These ingredients will determine the ﬁnal color, its lightness or

30

darkness. All of the labels read one of the following phrases; “individual shades do not
contain every ingredient” or “may contain.”

There was one active ingredient that was listed as may or may not being present
in seven of the dyes however, it was not listed at all on the label of Preference
Smoldering Red. This ingredient was propylene glycol.

Propylene glycol is a petroleum derivative that is formulated into hair care
products and serves as a humectant or a wetting agent. The role of a humectant is that it
retains the moisture content of cosmetic products by preventing the escape of moisture or
water. This substance causes the hair to swell due to the excess amount of water as a
result the amount of dye intermediates and precursors can not enter to develop. Hence,
after all things considered, hair ﬁbers dyed with Preference Smoldering Red required less
dye for consistency to be achieved. In addition to Feria All-Out Red Copper-Red and
Feria Glowing Red-Hot Red, which may not have contained this ingredient. All three of
these dyes that were previously listed only required 1 ml of dye for consistency to be
achieved.

The secondary purpose of this study was to determine if there were any
signiﬁcant spectral differences in hair bleaching formulas. A qualitative analysis of the
bleaching formulas illustrated that there were not any signiﬁcant spectral differences,
therefore for the most part their chemical composition is similar.

When the hair ﬁbers were bleached in this study, the pigments in the hairs were
eliminated. This is visible when looking at the overlay of spectra in Figure 24. There is

no deﬁnite shape to any of the spectra.

31

CHAPTER FIVE: CONCLUSION AND FUTURE RESEARCH

Conclusion

This study demonstrated that visible transmission microspectrophotometry was
shown to be an effective method for the examination of pigment distribution throughout
cosmetically treated hair ﬁbers. The presence of artiﬁcial treatments such as dyes and
bleaches can be identiﬁed through microscopical examination. The microscopic
association of bleached hairs is rather difﬁcult because these hair types have less
distinctive pigment patterns. Head hairs grow at a rate of one centimeter per month. The
approximate time of this treatment can be determined by measuring the length of the
untreated area of the hair.

Microspectrophotometry is a non-destructive method that provides useful
structural information and has the ability to accurately discriminate between similar dyes
using only a single hair ﬁber. This method does not provide conclusive forensic evidence,
but can serve as a screening test for hair identiﬁcation with other analytical and chemical
techniques such as gas chromatography and thin layer chromatography.

This study carried out by microspectrophotometry has revealed that it can serve
as a useful tool in to provide an objective rather than subjective information about color

in both the areas of forensic science and the cosmetic industry.

Future Research

Hair growth occurs both longitudinally and in diameter. It is well known from the
literature that the rate of hair growth varies from person to person. Hair grth is a
function of individual factors. Other factors such as age, gender, hair color, the presence

of drugs, hormones and nutrition can either accelerate or prolong hair growth. While

32

some of the factors may be or may not be known in an individual case, hair analysis from
a particular individual can be rather challenging.

Upon conclusion of this study, there are several possible research projects that
could derive from the present work. There is a need for research furthering the inclusion
of all racial hair types, since only Caucasian hair samples were used in this study. There
are striking and well documented differences between hair types of varying ethnic
origins, including Negroids, Caucasoids and Mongoloids. Hair from these groups differs
in numerous properties including maximum attainable length, ﬁber shape, thickness,
density, amino acid content, and tensile properties such as elasticity and percent
elongation at break. The effects of hair treatment depend on the characteristics of the
individual ﬁbers. The criteria required to perform racial hair analysis are summarized in

Table 6.

Table 6 - Racial Characteristics of Hair

 

 

 

 

Race Cross Diameter Pigmentation Cuticle Appearance
Section
Negroid Flat 60-90 pm Dense Thin Wavy
Caucasoid Oval 70—100 urn Even Medium Straight/wavy
Mongoloid Round 90-120 um Auburn Thick Straight

 

 

 

 

 

 

 

 

Before any permanent color can penetrate the hair shaft, the cuticle must be
opened so that the chemicals can get into the natural pigment molecules. When

comparing the racial characteristics of hair from Table 6, it is likely that the Caucasoid or

33

the Negroid hair types would always require less dye to achieve consistency than the
Mongoloid. The Mongoloid hair type possesses a thick cuticle and a larger cross section
as a result more dye molecules will be needed to penetrate the hair shaft and for the equal
distribution of the dye molecules throughout the entire hair strand. Additional research is
needed to draw a general conclusion about ethnic dependency.

As a continuation of the present research, different brands as well as colors of
permanent hair dyes could be analyzed to compare the amount of the dye that is need for
consistency to be achieved. These spectra of dye smears can be compiled to establish a
library for comparison of permanent hair dyes.

Additional research needs to be performed on developing
microspectrophotometry as a better discriminatory technique. Microspectrophotometry
was not able to conclusively distinguish between permanent hair dyes that had different
sub-brands.

Environmental factors have the ability to change the color of hair both physically
and chemically. When dyed or bleached hair is exposed to environmental factors such as
sun, water and air, it begins to oxidize and the dye pigments are released. The cosmetic
value of hair can be diminished through excessive exposure to the sun. Prolonged sun
exposure to hair is an example of photochemical change to the hair shaft. The UV light in
direct sunlight affects the cuticle in a similar way to bleach, by ultimately breaking down
the keratin protein in the hair.

Hair discoloration also occurs from both chlorinated and salt water. In dyed
blonde or bleached hairs, the chemicals in pool water tend to bind to the hair and cause

the hair ﬁbers to become green. Mineral deposits from salt water severely dehydrate hair.

34

The best way to minimize weathering damage is to use color safe shampoo and
deep conditioning treatments that contain keratin, jojoba oil and wheat germ. These
natural ingredients increase moisture and shine. After all things considered, two areas of
potential research are suggested: to determine what length of time is required for
cosmetically treated hair to become physically and chemically changed while being
exposed to these external factors and how well do these products work to minimize or

prevent damage from environmental exposure.

35

APPENDICES

36

APPENDIX I

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57

APPENDIX III

SPECTRA OF BLEACHED HAIR FIBERS

58

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59

APPENDIX IV

PHOTOGRAPH LIBRARY

60

 

Figure 25— Photo of Hair Fiber before Cosmetic Treatments
(Hair Fibers Used in Batches 1-14)

 

Figure 26 — Photo of Hair Fiber after Bleaching
(Hair Fibers Used in Batches 1-14)

61

 

Figure 27 - Photo of Hair Fiber from Batch 1 @ 1.0 ml of dye

 

Figure 28 — Photo of Hair Fiber from Batch 1 @ 1.5 ml of dye

62

 

Figure 30 — Photo of Hair Fiber Batch 3 @ 1.0 ml of dye

63

 

Figure 32 — Photo of Hair Fiber from Batch 4 @ 1.0 ml of dye

 

Figure 34 — Photo of Hair Fiber from Batch 5 @ 1.5 ml of dye

65

 

Figure 35 — Photo of Hair Fiber from Batch 6 @ 1.0 ml of dye

 

Figure 36 - Photo of Hair Fiber from Batch 7 @ 1.0 ml

 

Figure 37 — Photo of Hair Fiber from Batch 7 @ 1.5 m1 of dye

 

Figure 38 — Photo of Hair Fiber from Batch 8 @ 1.0 ml of dye

67

 

Figure 39 — Photo of Hair Fiber from Batch 8 @ 1.5 m1

 

Figure 40 — Photo of Hair Fiber from Batch 9

68

 

Photo of Hair Fiber from Batch 10

Figure 41

 

or from Batch 11

Fib

air

Figure 42 — Photo of H

69

 

Figure 44 - Photo of Hair Fiber from Batch 13

70

 

Figure 45 - Photo of Hair Fiber from Batch 14

71

APPENDIX V

INGREDIENTS

72

INGREDIENTS

Mamie—Ila

L’Oreal Excellence Red Penny

m-arninophenol, p-aminophenol, o-arninophenol, ammonium hydroxide, butoxydiglycol,
cocamide mipa, deceth-3, deceth-S, 2,4-diaminophenoxyethanol HCl, dicocodimonium
chloride, EDTA, erythorbic acid, fragrance, hydroxyethyl oleyl dirnonium chloride,
isopropyl alcohol, p-methylarninophenol sulfate, 2-methyl-5-hydroxyethylaminophenol,
2-methylresorcinol, p-phenylenediamine, propylene glycol, resorcinol, sodium
metabisulﬁte, thiolactic acid, water.

L’Oreal Feria All-Out Red Copper Red

L’Oreal F eria Double Intensity Auburn Red

L’Oreal Feria Glowing Red-Hot Red

alcohol denatured, 4-amino-2-hydroxytoluene, m-aminophenol, p-aminophenol, o-
aminophenol, ammonium acetate, ammonium hydroxide, ammonium thiolactate, 2,4-
diarninophenoxyethanol HCl, erythorbic acid, fragrance, hexylene glycol,
hydroxybenzomorpholine, 6-hydroxyindole, hydroxypropyl bis(n-hydroxyethyl-p-
phenylenediamine) HCl, 2-methyl-5-hydroxyethylaminophenol, p-methylaminophenol
sulfate, 2-methylresorcinol, n,n-bis(2-hydroxyethyl)-p-phenylenediamine sulfate, oleic
acid, oleyl alcohol, peg-2 oleamine, pentasodium pentetate, phenyl methyl pyrazolone,
p-phenylenediamine, polyglyceryl-2 oleyl ether, polyglyceryl-4 oleyl ether, propylene
glycol, resorcinol, sodium diethylaminopropyl, cocoaspartamide, sodium metabisulﬁte,
toluene-2,5-diamine, trideceth-2 carboxamide mea, water. Covered under US. Patents:
#5,114,429, #5,279,620, #5,900,028, #5,690,696 and #6,093,220. Individual shades do
not contain every ingredient.

L’Oreal Preference Red Penny

water, trideceth-2 carboxamide mea, propylene glycol, hexylene glycol, peg-2 oleamine,
polyglyceryl-2 oleyl ether, oleyl alcohol, alcohol denatured, ammonium hydroxide,
polyglyceryl-4 oleyl ether, oleic acid, sodium diethylaminopropyl, cocoaspartamide,
pentasodium pentetate, ammonium acetate, sodium metabisulﬁte, ﬁagrance, erythorbic
acid, ammonium thiolactate, thiolactic acid. May contain: p-phenylenediamine, 2-methyl-
5-hydroxyethylaminophenol, p-aminophenol, resorcinol, phenyl methyl pyrazolone, p-
methylaminophenol sulfate, 4-amino-2-hydroxytoluene, m-arninophenol, 2,4-
diaminophenoxyethanol HCl, 6-hydroxyindole, 2-methylresorcinol, hydroxypropyl bis(n-
hydroxyethyl-p-phenylenediamine) HCl, n,n-bis(2-hydroxyethyl)-p-phenylenediamine
sulfate, 2-amino-3 -hydroxypyridine, o-aminophenol, hydroxybenzomorpholine. Covered
under US. Patents: #5,279,620, #5,]14,429, #5,690,969 and #6,093,220.

L’Oreal Preference Smoldering Red

alcohol denatured, 2-amino-3-hydroxypyridine, 4-amino-2-hydroxytoluene, m-
aminophenol, p-aminophenol, p-phenylenediamine, ammonium acetate, ammonium

73

hydroxide, butoxydiglycol, 2,4-diaminophenoxyethanol hcl, ﬁagrance, erythorbic acid,
hydroxybenzomorpholine, 6-hydroxyindole, hydroxypropyl bis(n-hydroxyethyl-p-
phenylenediamine) HCl, p-methylarninophenol sulfate, 2-methyl-5-
hydroxyethylaminophenol, 2-methylresorcinol, n,n-bis(2-hydroxyethyl)-p-
phenylenediamine sulfate, oleic acid, oleyl alcohol, peg-2 tallow amine, pentasodium
pentetate, phenyl methyl pyrazolone, polyglyceryl-2 oleyl ether, polyglyceryl-4 oleyl
ether, propylene glycol, resorcinol, sodium diethylarninopropyl cocoaspartamide, sodium
metabisulﬁte, thiolactic acid, trideceth-2 carboxamide mea, water. Covered under US.
Patents: #5,] 14,429, #4,361,421 and #5,279,620. Every shade does not contain all of
these ingredients.

L’Oreal Preference Mega Reds Dark Intense Copper Red

L’Oreal Preference Mega Reds Medium Intense Red Copper

trideceth-2 carboxanride mea, water, propylene glycol, hexylene glycol, peg-2 oleamine,
polyglyceryl-4 oleyl ether, oleyl alcohol, alcohol denatured, ammonium hydroxide,
polyglyceryl-2 oleyl ether, oleic acid, sodium diethylarninopropyl cocoaspartamide,
pentasodium pentetate, ammonium acetate, sodium metabisulﬁte, fragrance, erythorbic
acid, ammonium thiolactate, thiolactic acid. May contain: p-phenylenediamine, 2-methyl-
5-hydroxyethylaminophenol, p—aminophenol, resorcinol, phenyl methyl pyrazolone, p-
methylanrinophenol sulfate, 4-amino-2-hydroxytoluene, m-arninophenol, 2,4-
diaminophenoxyethanol HCl, 6-hydroxyindole, 2-methylresorcinol, hydroxypropyl bis(n-
hydroxyethyl-p-phenylenediamine) HCl, n,n-bis(2-hydroxyethyl)-p-phenylenediamine
sulfate, 2-amino-3-hydroxypyridine, o-aminophenol, hydroxybenzomorpholine. Covered
under US. Patents: #5,279,620, #5,] 14,429, #5,690,969 and #6,093,220.

Bleach Ingedients

Wella Wellite Powder Lightener

potassium persulfate, sodium silicate, magnesium carbonate hydroxide, mineral oil,
ammonium persulfate, sodium stearate, xanthan gum, rice starch, algin, disodium EDTA,
silica.

Clairol Professional Kaleidocolors

potassium persulfate, sodium metasilicate, sodium stearate, silica, hydrated silica, sodium
lauryl sulfate, hydroxypropyl methylcellulose, sodium persulfate, ammonium persulfate,
disodium EDTA, aloe, barbadensis gel, fragrance, carbomer, dextrin, ultramarines, ext.
D&C violet no. 2.

Salon Care Quick White
potassium persulfate, sodium metasilicate, ammonium persulfate, sodium stearate, silica,
hydroxypropyl methylcellulose, aluminium stearate, EDTA, sodium lauryl sulfate.

L’Oreal Super Oreal Blane

potassium persulfate, sodium silicate, sodium persulfate, acrylates/c10-30 alkyl acrylate,
crosspolymer, urea, kaolin, magnesium stearate, ammonium chloride, diethylhexyl

74

sodium sulfosuccinate, vp/va copolymer, polydecene, sodium metasulﬁte, magnesium
peroxide, EDTA, titanium dioxide, cyamopsis tetragonoloba gum, sodium carboxymethyl
starch, sodium benzoate.

75

NOTES

 

Chapter One: Introduction

1 Robertson, J., and Aitken, C.G.G., “The Value of Microscopic Features in the
Examination of Human Head Hairs: Analysis of Comments Contained in
Questionnaire Returns,” Journal of Forensic Sciences, Vol. 31, No. 2, 1999, pp.
563-573.

2 Euring, M.B., “Visible Microscopial Spectrophotometry in the Forensic Sciences,”
Forensic Science Handbook, Vol. I, 2“d ed. (New Jersey: Prentice Hall, 2002), pp.
322-3 87.

3 www.kolors.com/pages/chemistry.html.

4 Savolainen, Peter and Lundeberg, Joakim., “Forensic Evidence Based on mtDNA
from Dog and Wolf Hairs,” Journal of Forensic Sciences, Vol. 44, No. 2, 1999,
pp. 77-81.

5 Benner, Bruce A., Jr., Goodpaster, John V., Degrasse, Jeffrey A., Tully, Lois A., Levin,
Barbara C., “Characterization of Surface Organic Components of Human Hair by
On-Line Supercritical Fluid Extraction-Gas Chromatography/Mass Spectrometry:
A Feasibility Study and Comparison with Htunan Identiﬁcation Using
Mitochondrial DNA Sequences,” Journal of Forensic Sciences, Vol. 48, No. 3,
2003,p.554.

6 Tanada, N., Kashimura, S., Kageura, M., Hara, K., “Practical GC/MS Analysis of
Oxidative Dye Components in Hair Fiber as a Forensic Investigative Procedure,”
Journal of Forensic Sciences, Vol. 44, No. 2, 1999, pp. 292-296.

7 www.kolors.com/pages/chemistry.html.

8’9 Wolfram, L.J., Hall, K., Hui, I., “The Mechanism of Hair Bleaching,” Journal of the
Society of Cosmetic Chemists, Vol. 21, 1970, pp. 875-900.

'0 “Types of Hair Bleaching: Hair Bleaching,” http:library.thinkquest.org/27034/hair.htrnl

” Skopp, G., Potsch, L., Moeller, M.R., “On Cosmetically Treated Hair — Aspects and
Pitfalls of Interpretation,” Forensic Science International, Vol. 84, 1997, pp.43-
52.

'2 “Types of Hair Bleaching: Hair Bleaching,” http:library.thinkquest.org/27034/hair.html

'3 Tanada, N., Kageura, M., Hara, K., Hieda, Y., Takamoto, M., Kashimura, S.,

“Demonstration of Oxidative Dyes on Human Hair,” Forensic Science
International, Vol. 64, 1994, pp.1-8.

76

Chapter 2: Instrumentation

'4 Olson, L.A., “Color Comparison in Questioned Document Examination Using
Microspectrophotometry,” Journal of Forensic Sciences, Vol. 31, No. 4,
Oct.1986, pp. 1330-1340.

'5’“ Euring, M.B., “Visible Microscopial Spectmphotometry in the Forensic Sciences,”
Forensic Science Handbook, Vol. I, 2"‘1 ed. (New Jersey: Prentice Hall, 2002), pp.
322-3 87

Chapter 3: Methods and Materials

'7 Euring, M.B., “Visible Microscopial Spectrophotometry in the Forensic Sciences,”
Forensic Science Handbook, Vol. 1, 2n ed. (New Jersey: Prentice Hall, 2002),
pp. 322-387.

Chapter 4: Results and Discussion

18 Corbett, J .F., “Chemistry of Hair Colorant Processes — Science as an Aid to

Formulation and development.” Journal of the Society of Cosmetic Chemists, Vol.
35, 1984, pp. 297-310.

'9 Corbett, J .F ., “Hair Coloring Processes.” Cosmetics & Toiletries, Vol. 106, 1991,
pp. 53-57.

77

BIBLIOGRAPHY

Bisbing, RE, “The Forensic Identiﬁcation and Association of Human Hair,” Forensic
Science Handbook, Vol. 1, 2nd ed. (New Jersey: Prentice Hall, 2002), pp. 389-428.

Benner, Bruce A., Jr., Goodpaster, John V., Degrasse, Jeffrey A., Tully, Lois A., Levin,
Barbara C., “Characterization of Surface Organic Components of Human Hair by
On-Line Supercritical Fluid Extraction-Gas Chromatography/Mass Spectrometry:
A Feasibility Study and Comparison with Human Identiﬁcation Using
Mitochondrial DNA Sequences,” Journal of Forensic Sciences, Vol. 48, No. 3,
2003,p.554.

Caspersson, T., “Methods for the Determination of the Absorption Spectra of Cell
Structures,” Transactions of the Royal Microscopical Society, 1940, pp. 8-25

Deedrick, D.W., “Hairs, Fibers, Crime and Evidence,” Forensic Science
Communications, Vol. 2, No.3, Jul. 2000,
www.fbi.gov/hq/lab/fsc/backissu/july2000/deedrick.htm

Euring, M.B., “Visible Microscopial Spectrophotometry in the Forensic Sciences,”
Forensic Science Handbook, Vol. 1, 2nd ed. (New Jersey: Prentice Hall, 2002), pp.
322-3 87.

Olson, L.A., “Color Comparison in Questioned Document Examination Using
Microspectrophotometry,” Journal of Forensic Sciences, Vol. 31, No. 4,
Oct.1986, pp. 1330-1340.

Pfeffferli, P.W., “Application of Microspectrophotometry in Document Examination,”
Forensic Science International, Vol. 23, No. 2,3, Nov/Dec. 1983, pp. 129—136.

Robertson, J., and Aitken, C.G.G., “The Value of Microscopic Features in the
Examination of Human Head Hairs: Analysis of Comments Contained in
Questionnaire Returns,” Journal of Forensic Sciences, Vol. 31, No. 2, 1999, pp.
563-573.

Skopp, G., Potsch, L., Moeller, M.R., “On Cosmetically Treated Hair — Aspects and
Pitfalls of Interpretation,” Forensic Science International, Vol. 84, 1997, pp.43-
52.

Suzuki, S., Suzuki, Y., Ohta, H., Sugita, R., Marumo, Y., “Microspectrophotometric
Discrimination of Single Fibres Dyed by Indigo and Its Derivatives Using
Ultraviolet-Visible Transmittance Spectra,” Science & Justice, Vol. 4], No. 2,
Apr./Jun. 2001, pp. 107-11].

78

Tanada, N., Kageura, M., Hara, K., Hieda, Y., Takamoto, M., Kashimura, S.,
“Demonstration of Oxidative Dyes on Human Hair,” Forensic Science
International, Vol. 64, 1994, pp.1-8.

Tanada, N., Kashimura, S., Kageura, M., Hara, K., “Practical GC/MS Analysis of
Oxidative Dye Components in Hair Fiber as a Forensic Investigative Procedure,”
Journal of Forensic Sciences, Vol. 44, No. 2, 1999, pp. 292-296.

Tucker, H.H., “The Coloring of Human Hair with Semipermanent Dyes,” Journal of the
Society of Cosmetic Chemists, Vol. 22, 197], pp. 379-398.

Wolfram, L.J., Hall, K., Hui, I., “The Mechanism of hair Bleaching,” Journal of the
Society of Cosmetic Chemists, Vol. 21, 1970, pp. 875-900.

79

   

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