Thursday, 27 October 2016
Friday, 14 October 2016
Spectrophotometer
Devices and mechanism
Every chemical compound absorbs, transmits, or reflects light (electromagnetic radiation) over a certain range of wavelength. Spectrophotometry is a measurement of how much a chemical substance absorbs or transmits. Spectrophotometry is widely used for quantitative analysis in various areas (e.g., chemistry, physics, biology, biochemistry, material and chemical engineering, clinical applications, industrial applications, etc). Any application that deals with chemical substances or materials can use this technique. In biochemistry, for example, it is used to determine enzyme-catalyzed reactions. In clinical applications, it is used to examine blood or tissues for clinical diagnosis. There are also several variations of the spectrophotometry such as atomic absorption spectrophotometry and atomic emission spectrophotometry.
A spectrophotometer is an instrument that measures the amount of photons (the intensity of light) absorbed after it passes through sample solution. With the spectrophotometer, the amount of a known chemical substance (concentrations) can also be determined by measuring the intensity of light detected. Depending on the range of wavelength of light source, it can be classified into two different types:
- UV-visible spectrophotometer: uses light over the ultraviolet range (185 - 400 nm) and visible range (400 - 700 nm) of electromagnetic radiation spectrum.
- IR spectrophotometer: uses light over the infrared range (700 - 15000 nm) of electromagnetic radiation spectrum.
In visible spectrophotometry, the absorption or the transmission of a certain substance can be determined by the observed color. For instance, a solution sample that absorbs light over all visible ranges (i.e., transmits none of visible wavelengths) appears black in theory. On the other hand, if all visible wavelengths are transmitted (i.e., absorbs nothing), the solution sample appears white. If a solution sample absorbs red light (~700 nm), it appears green because green is the complementary color of red. Visible spectrophotometers, in practice, use a prism to narrow down a certain range of wavelength (to filter out other wavelengths) so that the particular beam of light is passed through a solution sample.
Figure 1 illustrates the basic structure of spectrophotometers. It consists of a light source, a collimator, a monochromator, a wavelength selector, a cuvette for sample solution, a photoelectric detector, and a digital display or a meter. Detailed mechanism is described below.
A spectrophotometer, in general, consists of two devices; a spectrometer and a photometer. A spectrometer is a device that produces, typically disperses and measures light. A photometer indicates the photoelectric detector that measures the intensity of light.
- Spectrometer: It produces a desired range of wavelength of light. First a collimator (lens) transmits a straight beam of light (photons) that passes through a monochromator (prism) to split it into several component wavelengths (spectrum). Then a wavelength selector (slit) transmits only the desired wavelengths, as shown in Figure 1.
- Photometer: After the desired range of wavelength of light passes through the solution of a sample in cuvette, the photometer detects the amount of photons that is absorbed and then sends a signal to a galvanometer or a digital display, as illustrated in Figure 1.
Adams Chromatic Valence Color Space
Adams chromatic valence color spaces are a class of color spaces suggested by Elliot Quincy Adams.
Two important Adams chromatic valence spaces are CIELUV and Hunter Lab.
Chromatic value/valence spaces are notable for incorporating the opponent process model, and the empirically-determined 2½ factor in the red/green vs. blue/yellow chromaticity components (such as in CIELAB).
In 1942, Adams suggested chromatic value color
spaces. Chromatic
value, or chromance, refers to the intensity of the opponent
process responses, and is derived from Adams' theory of color
vision.
A chromatic value space consists of three components:
·
{\displaystyle
V_{Y},}VY the Munsell-Sloan-Godlove
value function: {\displaystyle
V_{Y}^{2}=1.4742Y-0.004743Y^{2}}(VY)2 = 1.4742Y –
0.004743Y2
·
{\displaystyle
V_{X}-V_{Y}}Vx – Vy, the red-green chromaticity dimension, where {\displaystyle V_{X}}Vx is
the value function applied to {\displaystyle
(y_{n}/x_{n})X}(yn/xn)X instead of Y
·
{\displaystyle
V_{Z}-V_{Y}}VZ - VY, the
blue-yellow chromaticity dimension, where {\displaystyle V_{Z}}VZ is the value
function applied to {\displaystyle
(y_{n}/z_{n})Z}(yn/zn)Z instead of Y
A chromatic value diagram is a plot of {\displaystyle V_{X}-V_{Y}}VX
- VY (horizontal axis) against {\displaystyle 0.4(V_{Z}-V_{Y})}0.4(VZ – VY) (vertical
axis). The 2½ scale factor is intended to make radial distance from the white point correlate
with the Munsell chroma along any one hue
radius (i.e., to make the diagram perceptually uniform). For achromatic surfaces, {\displaystyle
(y_{n}/x_{n})X=Y=(y_{n}/z_{n})Z}(yn/xn)X = Y = (yn/zn)Z
and hence {\displaystyle
V_{X}-V_{Y}=0}, {\displaystyle
V_{Z}-V_{Y}=0}VX – VY = 0, VZ – VY
= 0. In other words, the white point is at the origin.
Constant differences along the chroma dimension did not appear different
by a corresponding amount, so Adams proposed a new class of spaces, which he
termed chromaticvalence. These spaces have "nearly equal radial
distances for equal changes in Munsell chroma".
Kubelka-Munk equations:
When light is fall on some sample, some part of the light reflected, some
are absorbed and some are scattered. If the sample is having opacity more than
70% then as per Kubelka Munk (established in 1931) following is the relation
between reflectance, scattering and absorption of light:
K/S= (1 – 0.01R)2/2(0.01R)
The mathematical basis for all color matching software is the Kubelka-Munk
series of equations. These equations state that for opaque samples such as
textile materials, the ratio of total light absorbed and scattered by a mixture
of dyes is equal to the sum of the ratios of light absorbed and scattered by
the dyes measured separately. Where absorption is defined as "K" and
scattering is defined as "S", Kubelka-Munk states that _:
(K/S) mixture = (K/S) dye 1 + (K/S) dye 2 + (K/S) dye 3 + ...
K/S is not a readily measurable quantity, but it can be calculated from the
reflectance of a sample -- "R" -- by the Kubelka-Munk equation that
states
K/S= (1-R)2/2R
K/S value is proportional to dye concentration in the substrate
K/S=kC
Where k is constant and C is concentration of
dyes or colorant
Delta E Differences and Tolerances.
The difference between two colour samples is often
expressed as Delta E, also called DE, or ΔE. 'Δ' is the Greek letter for 'D'. This
can be used in quality control to show whether a dyed or printed sample, such
as a colour swatch or proof, is in tolerance with a reference sample or
industry standard. The difference between the L*, a* and b* values of the
reference and sample will be shown as Delta E (ΔE).
The resulting Delta E number will show how far apart visually the two samples
are in the colour 'sphere'
CIE L*C*h°
This is possibly a little easier to
comprehend than the Lab colour
space, with which it shares several features. It is more correctly known
as L*c*h*. Essentially it is in the form of a sphere. There are three
axes; L* , c* and h°.
The L* axis represents Lightness.
This is vertical; from 0, which has no lightness (i.e. absolute black), at the
bottom; through 50 in the middle, to 100 which is maximum lightness (i.e.
absolute white) at the top.
The c* axis
represents Chroma or 'saturation'. This ranges from 0 at
the centre of the circle, which is completely unsaturated (i.e. a neutral grey,
black or white) to 100 or more at the edge of the circle for very high Chroma
(saturation) or 'colour purity'.
The h* axis represents Hue. If we
take a horizontal slice through the centre, cutting the 'sphere' ('apple') in
half, we see a coloured circle. Around the edge of the circle we see every
possible saturated colour, or Hue. This circular axis is known as h° for Hue. The
units are in the form of degrees (or angles), ranging from 0° (red) through 90°
(yellow), 180° (green), 270° (blue) and back to 0°.
The Lch colour model is very useful for
retouching images in a colour managed workflow, using high-end editing
applications. Lch is device-independent.
CIE
XYZ:
The basic CIE colour space, or colour
model, is based on a 'Standard Observer and 'Standard Illuminants' (D50, D65,
etc.). This is a numerical model of colour sensitivity based on research commenced
in the 1920s on a sample of people with normal colour vision. It is a
'universal colour space' representing the colour spectrum visible to the
'average human'. The light-sensitive retina at the back of the eye has three
types of receptors near the centre, known as cones.
They are sensitive to the three primaries, 'red, green and blue'. The CIE XYZ tristimulus values are assigned to the red,
green and blue curves respectively. These approximate to the cones in the eye. The relative
response of each is plotted on a diagram against the wavelength in nanometers.
The eye also has rods, outside of the
retina's centre, which are sensitive to low-wavelength light and which only
operate at low levels of illumination. There are two axes: vertical and
horizontal.
The vertical axis represents Relative Response 0 -
2.0 (shown here) or Reflective Intensity 0 - 120% (not shown).
The horizontal axis represents
Wavelength in nanometers, usually from about 380 to about 720nm.
It should be emphasized that this is a
'device-independent' colour space in which each primary colour (X,Y,Z) is always constant,
unlike RGB which varies with every individual device (monitor, scanner,
camera, etc.). XYZ is typically used to report the spectral response of a
sample measured by a colorimeter or a spectrophotometer. A colorimeter may
contain as few as three sensors, one each for red, green and blue, (or X, Y and
Z), and will typically be used for display calibration and profiling. A
spectrophotometer will report the entire spectral response at frequent
intervals along the spectrum, say every 10 nanometres, and will typically be
used to measure printed sheets to control a press or create an ICC profile.
While CIE XYZ is used to report colour
from measuring instruments, it is not so useful for humans to describe colour.
Another use is as the Profile Connection Space (PCS)
within an ICC profile, where it may be used instead of CIE Lab.
You may notice that the Y ('green
curve') covers the widest wavelength. This corresponds to the overall human
visual response to all colours, or lightness. It is therefore also used to
indicate luminance ('lightness').
CIE L*a*b*: In such case the vertical L* axis represents Lightness,
ranging from 0-100. The other (horizontal) axes are now represented
by a* and b*. These are at right angles to each other and cross
each other in the centre, which is neutral (grey, black or white). They are
based on the principal that a colour cannot be both red and green, or
blue and yellow.
The a* axis
is green at one extremity (represented by -a),
and red at the other (+a). The b* axis has blue at one end (-b), and yellow (+b) at
the other.
The centre of each axis is 0. A value of 0, or very low numbers of both a* and b* will describe a neutral or near neutral. In the case of paper, the white point in terms of a* and b* is usually carried through to the black, being gradually reduced towards '0'.
The centre of each axis is 0. A value of 0, or very low numbers of both a* and b* will describe a neutral or near neutral. In the case of paper, the white point in terms of a* and b* is usually carried through to the black, being gradually reduced towards '0'.
In theory there are no maximum values
of a* and b*, but in practice they are usually numbered from
-128 to +127 (256 levels).
The CIE Lab colour model encompasses
the entire spectrum, including colours outside of human vision. CIE Lab is
extensively used in many industries apart from printing and photography. Its
uses include providing exact colour specifications for paint (including
automotive, household, etc.), dyes (including textiles, plastics, etc.),
printing ink and paper.
CIE Color Systems
CIE Color Systems The CIE, or Commission Internationale de l’Eclairage
(translated as the International Commission on Illumination), is the body
responsible for international recommendations for photometry and colorimetry.
In 1931 the CIE standardized color order systems by specifying the light
source (or illuminants), the observer and the methodology used to derive values
for describing color.
The CIE system
characterizes colour by a luminance Y and two colour coordinates x and y which
specify the point on the chromatic diagram. This system offers more precision
in colour measurement than do the Munsell system because the parameters are
based on the spectral power distribution of the light emitted from a coloured
object and is factored by sensitivity curves which have been measured for the
human eye.
Based
on the fact that the human eye has three different types of color sensitive cones, the response of
the eye is best described in terms of three "tristimulus values".
However, once this is accomplished, it is found that any color can be expressed
in terms of the two color coordinates x and y.
The
colors which can be matched by combining a given set of three primary colors (such as the blue, green, and red) are
represented on the chromaticity diagram by a triangle joining the coordinates
for the three colors.
The diagram given below represents
the mapping of human color perception in terms of two CIE parameters x and y. The spectral colors are distributed around the edge of the
"color space" as shown, and that outline includes all of the
perceived hues and provides a
framework for investigating color.
The CIE Color Systems utilize three coordinates to locate a color in a
color space. These color spaces include:
• CIE XYZ
• CIE L*a*b*
• CIE L*C*h°
Scales for Measuring
Colour:
There are two
important scales for measuring colour. These are:
• Munsell
Scale
• CIE
colour system
Munsell Scale:
In 1905, artist Albert H.
Munsell originated a color ordering system — or color scale — which is still
used today. The Munsell System of Color Notation is significant from a
historical perspective because it’s based on human perception. Moreover, it was
devised before instrumentation was available for measuring and specifying color. This system
assigned numerical value to the three properties of the colour-Hue, Chroma and
value. The Munsell color
system match colors to a set of standard samples. The Munsell system divides hue into
100 equal divisions around a color circle and circle is distorted by assigning
a unit of radial distance to each perceptable difference in saturation (called units of chroma). Since there
are more perceptable differences for some hues, the figure will bulge outward
to 18 values for some hues compared to only 10 for another. Perpendicular to
the plane formed by hue and saturation is the brightness
scale divided into a scale of "value" from 0 (black) to 10 (white). A
point in the color space so defined is specified by hue, value, and chroma in
the form H V/C.
The MUNSELL system is a collection of
color samples for comparison, with adjacent samples based upon equal perceived
differences in color.
Munsell saw that full
chroma for individual hues might be achieved at very different places in the
color sphere. For example, the fullest chroma for hue 5RP (red-purple) is
achieved at 5/26.
Another color such as
10YR (yellowish yellow-red) has a much shorter chroma axis and reaches fullest
chroma at 7/10 and 6/10:
Instrumentation for colour measurement:
As colour is perception, it
cannot be directly measured; however we can measure and subsequently calculate
certain factors which are responsible for producing this sensation of color.
The quantification of the color properties of textile materials is of great
economic value in industry and instruments are employed to some degree in
almost every textile operation involved in textile coloration.
Color instrumentation
has experienced a tremendous advancement in technology during the past few
decades. The first devices for measuring color were absorptiometers which were
used to determine by visual inspection whether two solutions were of equal
color. This is very similar to holding two glass cylinders of dye solution up
to a light and judging whether they are of equal strength and shade, except
that the absorptiometer provided a method of adjusting the thickness or path
width so that this change in width could be read from a scale. In measuring
reflected light from opaque materials such as textiles, the first instruments
were reflectometers developed around 1915-1920.
Perception of colour: It involves a series of events which are interdisciplinary in nature.
Perception of colours includes source of light, object that is illuminated and
eye and brain that perceive the colour.
Object
Metamerism: It is a phenomenon observed when two specimens appears to have the similar
colour under one set of viewing conditions, but different under another. The
change in viewing conditions refers to change in source, observer or geometry
of the observation.
Types of metamerism
•
illuminant
metamerism
•
observer
metamerism
•
geometry
metamerism
Illuminant metamerism
When the colour of two specimens matches under one
illuminant but not with another illuminant, it is said to be illuminant
metamerism.
Observer metamerism
When colour of two objects appears to match to one
observer but not to the other, it is said to be observer metamerism. In this
case observer may not be colour blind but the spectral sensitivity of colour
receptors of one observer may be slightly different than the other.
Geometry metamerism
Two objects which match in one arrangement of
illumination, sample and observer may mismatch by altering the positions.
MEASUREMENT
OF COLOUR
Colour is perception and sensation
experienced caused by light reflected from or transmitted through objects. If
you have some coloured object, it means it has
certain reflectance characteristics—the patterns of light wavelengths that are
reflected and absorbed—that are physical properties of object. However, colour
is our subjective perception of the wavelengths of light that end up bouncing
off the object and onto our retina. Photoreceptors in the retina begin the
process by selectively responding to different wavelengths. A single type of
photoreceptor alone cannot accomplish color vision since it cannot distinguish
between enough wavelengths. We
overcome this difficulty by using three different types of cone photoreceptors
to code for color, a concept known as trichromacy. One of the key observations
in developing the trichromatic theory of color vision was that lights can be
added together to form mixtures that look identical to other, single light
wavelengths. Another activity illustrates additive and other forms of color mixing.
Human Visual System
The
human visual system consists of two functional parts, the eye and (part of the)
brain. The brain does all of the complex image processing, while the eye
functions as the biological equivalent of a camera.
What
our eyes perceive of a scene is determined by the light rays emitted or reflected
from that scene. When these light rays are strong enough (have enough energy),
and are within the right range of the electromagnetic spectrum (about 300 to
700 nm), the healthy eye will react to such a ray by sending an electric signal
to the brain through the optic nerve. When a light ray hits the eye, it will
first pass through the cornea, then subsequently through the aqueous humor, the
iris, the lens, and the vitreous humor before finally reaching the retina. The
cornea is a transparent protective layer, which acts as a lens and refracts the
light. The iris forms a round aperture that can vary in size and so determines
the amount of light that can pass through. Under dark circumstances the iris is
wide open, letting through as much light as possible. In normal daylight, the
iris constricts to a small hole. The lens can vary its shape to focus the
perceived image onto the retina. In the retina, the light rays are detected and
converted to electrical signals by photoreceptors. The eye has two types of
photoreceptors: rods and cones, named after their approximate shape. The rods
are abundant, about 100 million in a human eye, and spread evenly about the
retina, except at the fovea, where there are almost none. The fovea is the area
of the retina where our vision is sharpest. There are much fewer cones, about 6
to 7 million, which are mainly located around the fovea, but can be found in a
low density in the entire retina. No photoreceptors are found at the point where
the optic nerve attaches to the eye (the so-called blind spot), so we cannot
perceive anything there. Since rods are more responsive to light than cones we
can identify three types of vision, depending on the amount of light that
reaches the eye. Under dark circumstances, practically only the rods are
active. Since rods cannot discriminate colors, we perceive only shades of grey.
We call this scotopic or night vision. Under daylight circumstances, the cones
are most active, and we experience photopic or day vision. In dimly lighted
circumstances there is an intermediate stage where both rods and cones are
active called mesopic vision. We are able to distinguish colors because there
are three distinct types of cones, each sensitive to a different band of the
electromagnetic spectrum.
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