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Color analysis of textile fibers by microspectrophotometry

Can Hu, Hongcheng Mei, Hongling Guo, Jun Zhu

PII: S2468-1709(20)30009-6
DOI: https://doi.org/10.1016/j.forc.2020.100221
Reference: FORC 100221

To appear in: Forensic Chemistry

Received Date: 17 June 2019

Revised Date: 10 September 2019
Accepted Date: 24 January 2020

Please cite this article as: C. Hu, H. Mei, H. Guo, J. Zhu, Color analysis of textile fibers by microspectrophotometry,
Forensic Chemistry (2020), doi: https://doi.org/10.1016/j.forc.2020.100221

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 Color analysis of textile fibers by microspectrophotometry

Can Hu, Hongcheng Mei, Hongling Guo, Jun Zhu*

Insititute of Forensic Science, Ministry of Public Security, PRC

Address correspondence to Jun Zhu, Insititute of Forensic Science, Ministry of Public

Security of China, No. 17 Muxidi nanli, Xicheng District, Beijing, PRC. E-mail:
zhujun001cn@126.com

Color analysis of textile fibers by microspectrophotometry

Abstract: Textile fibers are an important form of trace evidence, and color is a

pivotal characteristic of fiber evidence. The analysis of fiber color is crucial for

the reliable association or discrimination of fiber evidence. The

microspectrophotometry (MSP) technique is fast and non-destructive, and it

enables the measurement of color from small samples, which makes MSP an

ideal tool for color comparison of fiber evidence. We provide a review of the

advances made in recent years in color analysis of textile fibers using MSP.

The principle of MSP as a color comparison technique is introduced. The

increasing use of chemometric and statistical methods for the differentiation of

MSP spectra from dyed fibers are also discussed.

Keywords: Color; Microspectrophotometry; Textile fibers; Chemometric

Introduction

Textile fibers are an important form of trace evidence from crime scenes. The
transfer of textile fibers can link people, tools, and crime scenes by inferring contact

between two individuals or between individuals and objects. Color is a pivotal

appearance characteristic of fiber evidence that reflects the dyes and pigments applied

on the textile [1]. For many fibers, such as cotton, the only characteristic that can be

reliably used for discrimination of samples is its color. It is necessary to build

instrumental methods to compare fiber colors in forensic science. The color difference

between fiber evidence may be very small and undistinguishable by the naked eye.

Additionally, objective, high quality, and demonstrable data regarding the color are

indispensable in a trial.

Numerous methods have been developed for fiber color analysis, including

spectroscopy methods (spectrophotometry [2-4], infrared spectroscopy [5,6], Raman

spectroscopy [7,8], chromatographic methods (thin layer chromatography (TLC))

[9,10], high performance liquid chromatography (HPLC) [11,12], capillary

electrophoresis (CE) [13,14], and mass spectrometry (MS) methods [15,16]. Those

methods have been summarized and discussed in 2009 [17]. Spectroscopy methods are

frequently used because they are fast, non-destructive, and require little sample

preparation, unlike other methods that require extraction of the dye. Considering that

fibers recovered from crime scenes are often tiny, spectroscopy methods that are able

to analyze small samples are needed.

Microspectrophotometry (MSP) is a combination of microscopy and

spectrometry. It enables the measurement of a spectrum of a very small sample

(approximately 100 μm2) and examination of its morphology. The MSP technique is
fast and non-destructive, which makes it widely used in different fields. In life science,

MSP could be applied to determine biological staining [18,19], and the stained DNA

could be quantified by MSP. In the field of jewelry identification, MSP could be

employed for gem color analysis [20]. In forestry science, MSP has been applied to

locate lignin and other components in plant cell walls [21,22], and the MSP method

was found to be ideally suited to the study of the subcellular distribution in plant cell

walls. In forensic science, MSP has been widely applied in the color examination of

evidence, such as paints [23,24], inks [25], hair [26], and soil [27].

In 1868, Henry Clifton Sorby and John Browning combined a microscope and a

direct vision spectroscope, which pioneered a new scientific technology:

microspectroscopy [28]. This new instrument was applied to study the colors of a wide

range of natural pigments in biological materials. The microspectral study of blood led

to the first forensic applications. Since then, much progress has been made, and many

new detection targets have been published. In 2012, Janina Zieba-Palus et al. [29]

introduced the application of microspectrophotometry for comparative examination of

forensic evidence, such as paint chips, single fibers, and inks on questionable

documents. In 2009, S.Walbridge-Jones discussed the strengths and limitations of MSP

fortextile fiber color measurement [30]. Since then, no review article has been

published that summarizes the latest developments in color analysis of textile fibers by

MSP.

The aim of this review is to provide an outline of recent research advances in

fiber color comparison using MSP. A selected number of publications are reviewed
with the aim to present the most important detection strategies and applications. The

principle of MSP as a color comparison technique is introduced. The advantages and

disadvantages of the developed method and the requirement for further research and

development are also discussed.

The principle of color measurement by MSP

Human eyes can detect light in the range of 380-780nm, a portion of the

electromagnetic spectrum known as visible light, which we understand as color. Light

at the lower end of the visible spectrum has a longer wavelength of about 740 nm, it is

seen as red. Violet at the upper end of the spectrum having a wavelength of about 380

nm, and green is in the middle (Fig.1). The color of a sample depends on three factors:

the characteristics of the sample, the light source, and the color response of the eye [31].

Different substances with diverse chemical structures have various reflecting power and

absorbency to different wavelengths of light, which produce different optical spectra

[32,33]. The incident illumination and angle can also influence the spectrum as well as

the color of the object. The color response of the eyes is based upon the varying

sensitivity of cone cells; thus, visual descriptions of color are very subjective.
 Fig. 1 The relationship between wavelength, frequency and color.

According to the color ordering system proposed by the Commission

International de l’Eclairage (CIE) of 1931, each shade of color can be obtained by

mixing three primary colors (red, green, and blue) in appropriate proportions. The

amounts of the three primary colors contained in each color is generally referred to as

a tristimulus value, which is represented by X, Y, and Z, respectively [34]. The

proportion of red, blue, and green colors in a sample is represented by chromaticity

X Y Z
coordinates x, y, and z, i.e., x = X + Y + Z; y = X + Y + Z; z = X + Y + Z; x + y + z = 1.

We can convert x, y, and z into chromaticity values of L*, a*, and b*, and obtain

a set of L1*, a1*, and b1* for colored object 1, a set of L2*, a2*, and b2*for colored object

2. The color difference between the two objects is ∆E = (∆L ∗ )2 + (∆a ∗ )2 + (∆b ∗ )2,

where ΔL*=L1*- L2*, Δa*=a1*- a2*, and Δb*=b1*- b2*.

MSP can measure and produce spectral curves of light energy as the light is

transmitted, absorbed, or reflected by a sample. Figure 2 shows a diagram of the optical

path for MSP. Incident white light illumination allows for reflectance

microspectroscopy from the deep UV to the near IR. Incident illumination at

wavelengths in the excitation range of photoluminescent samples can also be used for

fluorescence or phosphorescence microspectroscopy. The spectrum obtained by MSP

can also be converted to chromaticity values (L*, a*, and b*), and thus, objects that have

different colors can be identified by their ΔE. The detected data eliminates the

subjectivity associated with human description of color. Thus, MSP is a valuable tool

for comparing colors.

Holographic CCD
Grating detector

Digital
Aperture
imager

Reflectance
Beamsplitter
Light source
Imaging
Optics
Sample

Focusing
Optics
Transmission
Mirror Light source

Fig. 2 Schematic diagram of the optical path for MSP.

Color measurement of textile fibers by MSP

During the process of fiber analysis, the following problems are usually

encountered: 1) the choice of transmission or reflection mode; 2) the selection of

ultraviolet (UV), visible (Vis), or a fluorescent light source; or 3) the analysis of the
results (spectra and chromaticity values).

Fibers are typically analyzed using the transmission mode. The application of the

transmission mode of MSP in fiber comparison requires special sample preparation, i.e.,

embedding in a mounting medium (such as glycerin) on glass/quartz microscope slides.

Quartz slides are used for UV MSP detection because quartz is transparent to UV light.

Furthermore, when applying MSP in the UV range, measurements can be taken only in

the transmittance mode.

UV, Vis, and fluorescence MSP have all been employed to analyze fibers [35-

37]. Although the Vis region (400–760 nm) is most frequently used, UV-Vis (200–760

nm) and fluorescence MSP can provide more information and better discrimination.

Research shows that Dyed fibres could be analysed under glass mount by UV-VIS

microspectrophotometry from 320 nm to 770 nm. For fibre types with little or no

polymeric UV spectral interference, additional spectral information could be obtained

from UV-VIS analysis on quartz slides, measured from 240 nm to 770 nm [38]. It's

worth noting that the fluorescence MSP is not applicable to all dyestuffs, and the CIE

system should be modified to code fluorescent colors. Moreover, the fluorescence

intensity of some dyestuffs may decrease with time [37].

There are two types of data obtained by MSP (Fig. 1), spectra and CIE

chromaticity values (L, a, and b). Spectra obtained from MSP are often normalized and

visually compared. Mathematical calculations can also be performed before

comparison. Wiggins et al. [39] calculated the first derivative of the absorbance spectra

for fiber analysis. When the absorbance spectra of some fibers were broad and
featureless, the first derivative spectra could provide more points of comparison to

facilitate discrimination. However, the first derivative spectra may show differences

between fibers that match. A representative control sample is hence essential. And the

first derivative data should never be considered in isolation from the original

absorbance spectrum. The chromaticity value obtained from MSP allows for

quantitative descriptions of fiber color. The use of the colorimetry value for fiber color

measurements was exhaustively discussed by Adolf et al. [36] Combined with the

chemometric method, it can improve the discriminating power of MSP when analyzing

the results, which will be described in detail later.

Fig. 1 Color measurement of a fiber by MSP. The morphology (A), transmission spectrum

(B), and chromaticity value (C) of the fiber.

The application of MSP to color analysis of textile fibers

The examination of fibers color is indispensable for reliable fiber identification in

case investigations. In 1986, Laing et al. [40] reported the primarily use of MSP to

discriminate visually identical fibers based on their visible absorption spectra. Since

then, the application of MSP to fiber analysis has emerged.

The discrimination power toward different colors

To quantitatively descript the discrimination power of MSP, a calculation method

was established. The number of unique pair-wise comparisons within a group of

samples is:

𝑐 = (𝑥−1)/2

Where c is the number of unique comparisons and x is the number of samples in

the group. The number of discriminated pairs of samples divided by c gives the

discriminatory power. It is notable that the calculated discriminatory power is

dependent on the size and diversity of the samples themselves.

The discrimination power of MSP toward different color fiber samples has been

compared. Cotton fibers of a similar color have been discriminated quite well by MSP,

and the discrimination degree is highest for blue, followed by red cotton fibers [41].

The same results were obtained when UV MSP was used to differentiate cotton samples:

the blue samples had the best distinction [42,43]. Thus, 100 non-denim blue cotton

fibers, including t-shirts, trousers, and jumpers, were subjected to analysis by both the

Vis and UV range via MSP [44]. The discriminating power of visible range MSP was

calculated to be 0.89 for ‘mid-blue’ garments and 0.87 for ‘dark blue’ garments.

Extending MSP into the UV range increased the discrimination by 7%, giving a

discriminating power of 0.96 for both mid and dark blue cotton fibers. Based on these

good discrimination powers, fiber samples of the same generic type having the same

subjective color, which are defined as “blocks of color” [45], were studied using MSP,

and subtypes of orange, green, blue, and red cotton fibers have been distinguished
according to their spectral features, particularly in the UV range [45,46]. A high

discriminating power (0.9985 for blue cotton fibers, and 0.9995 for red cotton fibers)

was obtained using a combination of light microscopy, fluorescence microscopy and

UV/VIS MSP.

However, when fibers are very lightly or very darkly dyed, the color

differentiation according to MSP may be difficult [36]. This may not be the case when

differentiating sulfur, reactive, and direct dyes according to the MSP measurements

results of numerous black cottons [47] (Grieve et al. 2001). Uncolored fibers are

difficult to analysis using MSP. However, they can fluoresce when irradiated with UV

light owing to the presence of fluorescent brighteners. A microspectrophotometer was

converted into a microspectrofluorimeter for the discrimination of fluorescent

brighteners and fluorescent red dyestuffs on single fibers [48].

The analysis of different classes of dyes

Although reactive dyes are widely used, it is difficult to extract them because of

their covalent bonds with fibers; thus, the analysis of colored fibers with reactive dyes

is of great concern [49]. Intra-sample and inter-sample variation as well as a dichroism

effect in the case of a cotton fibers dyed with reactive dye have been observed. No

correlation exists between color uniformity in cotton fabrics and the concentration of

the reactive dye. Cotton fibers dyed with vat dyes, such as indigo and indigo derivatives,

have also been studied by UV-VIS MSP [50]. Wool and polyamide fibers dyed with

analogous metal complex dyes have been analyzed by UV-Vis MSP [51]. Single fibers

selected from 15 woolen and 15 polyamide fabrics that were dyed with analogous
gryfalan dyes have been studied. Combined with chemometric treatment, the results for

the fibers provided good discrimination.

Other applications

Besides the discrimination of similar colors, other applications have also been

reported. Rees Powell et al. [52] developed a database of textile fiber MSP spectra.

Over 4,400 fibers were recovered and grouped. The database, which was designed to

compare MSP spectra using a modified Pearson method of correlation, currently

contains over 20,000 normalized and first derivative spectra of casework, validation,

and reference textile fibers. Mary Eng et al. [53] utilized UV-VIS MSP to analyze

metameric samples which were created using different coloring agents or different

relative concentrations of the coloring agents. However, only the spectra between 350

and 800 nm were recorded, if the wavelength could be expand to 200 nm, the sample

could be differentiated better. Forster et al. [54] employed MSP to examine the

photofading of a fiber dye. MSP is generally regarded as a nondestructive method, and

sometimes photofading of fiber color during analysis can occur. A visible reduction in

absorbance was observed for cotton fabric dyed with blue, yellow, and red direct dyes

after 30 min of MSP examination. Microscopy, MSP, and TLC have also been

employed for comparison of the dyes to examine the batch variation [55]. MSP has also

been used to analyze the dye transport phenomena in Hercosett-treated wool [56]. A

capillary microspectrophotometry (cMSP) method was built to obtain reproducible

spectra of the dyes from heterogeneous fibers [57]. Dyes were extracted and

subsequently analyzed in a borosilicate glass flat microcapillary cell by MSP. The

method can hence guarantee the reproducibility of results.

Combination with other methods

The use of MSP together with TLC is a long-standing method used for fiber

examination [55]. MSP is generally employed as a complementary technique to TLC,

and in some circumstances, MSP can distinguish fibers that are not discriminated by

TLC [36]. If the concentration of the dye in the fiber is not sufficient for TLC or a non-

destruction requirement exists for the fiber, MSP would be more appropriate than TLC.

The importance of TLC and MSP in the analysis of reactively dyed cotton fibers has

been described [58,59]. MSP in combination with TLC has also been used to compare

fiber dyes from different batches [60].

Raman spectroscopy allows for the in situ analysis of fibers. The technique is fast

and almost non-destructive, and no (or very minimal) sample preparation is needed [61].

It is possible to detect the presence of a dye in a single fiber with concentrations below

those that are detectable by MSP; thus, Raman spectroscopy has also been used in

combination with MSP for fiber color analysis. Massonnet et al. [62] employed Raman

spectroscopy and MSP to analysis reactive dyes on cotton fibers. The detection limit of

Raman was found to depend both on the chemical composition of the dye itself and on

the analytical conditions, particularly the laser wavelength. For dye mixtures, Raman

spectroscopy is more sensitive compared with MSP. A binary dye mixtures was

analyzed in this paper, but Raman is limited when come across a more complex mixed

dyes.

Lepot et al. [63] applied Raman spectroscopy to forensic fiber cases. The
mounted fibers from MSP detection were directly examined using Raman spectroscopy,

and the spectra showed a small amount of interference from the mounting resin and

glass. Raman spectroscopy is therefore a powerful method that can be applied in routine

fiber analysis following MSP measurements.

Markstrom et al. [64] utilized MSP in combination with a liquid crystal tunable

filter to analysis single textile fiber. The spectrum compared well with results from a

conventional MSP. Some of its advantages include rapid and simple sample preparation

and easy comparison of multiple fibers at the same time. The flexibility of the approach

is an advantage over most commercial MSP. Once the image is taken, numerous regions

of interest can be compared for accuracy. But to realize this function, the instrument

needs to be specially built.

Applying chemometric and statistical methods to MSP data can provide valuable

information for fiber analysis. Compared with visual examination, chemometric

methods identify patterns and groupings from large complex datasets more accurately

[65]. The use of a statistical approach can also make the analysis much less subjective.

Eric et al. [66] reported MSP analysis of yellow polyester fiber by agglomerative

hierarchical clustering (AHC), principal component analysis (PCA), and discriminant

analysis (DA). A good discrimination was obtained for fibers dyed with different

concentrations of the same dye. PCA, DA and Fisher’s exact test were employed to

compare questioned and known fibers after MSP analysis [67] with 118 out of 121 fiber

comparisons correctly classified. Although the use of statistical methods with MSP

shows great potential for fibers distinguishes, the statistical methods are not infallible
considering the instrumental variability and the heterogeneity of the sample.

Conclusions

The MSP method does not require difficult sample preparation, it is quick and non-

destructive, and its discriminating power is good. Therefore, MSP is a central method

used for color and dye examination of fibers. To improve the discrimination power of

MSP, more attention should be paid to the UV region of the spectrum. Chemometric

methods applied to MSP data analysis could also enhance the reliability of conclusions.

Increased reliance on mathematical techniques for comparing MSP data is a clear trend

in forensic science. A standard, statistical method needs to be established for the

comparison of MSP spectra data. Owing to the insufficient sensitivity, MSP can only

be used as a preliminary screening method for fiber examination in some cases. With

the fast development of instrumentation technologies, some methods (such as Raman

and ambient mass spectrometry) complement MSP well.

Disclosure statement
The authors declare no conflicts of interest.

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Highlights
 The principle of colour measurement by microspectrophotometry(MSP) is presented.
 MSP methods for colour analysis of textile fibers are introduced.
 The applications of MSP in colour analysis of fibers are discussed.
 Combined with other method could increase the discrimination power of MSP.
 Chemometric methods applied to MSP enhanced the reliability of conclusions.