Open access peer-reviewed chapter

A Digital Image-Based Colorimetric Technique Use for Quantification of Green Active Pharmaceuticals Obtained from Natural Sources

Written By

Vitthal V. Chopade and Jayashri V. Chopade

Submitted: 31 August 2021 Reviewed: 26 October 2021 Published: 20 July 2022

DOI: 10.5772/intechopen.101398

From the Edited Volume


Edited by Ashis Kumar Samanta

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Colorimetry is the determination of colors, as name indicates. This method can use for to find out the concentration of compound (solute) in a colored solution in terms of chemical analysis (solvent). We frequently need to quantify the quantity of a specific component in a combination or the concentration of a solution during scientific activity. The trick is to determine the color differences between various combinations and their absolute values. This is more instructive and scientifically valuable than relying on subjective judgments like the color of the solution. Colorimetry is the measurement of colors, as the name implies. It is the measurement of the concentration of a certain compound (solute) in a colored solution in terms of chemical analysis (solvent). We frequently need to quantify the quantity of a specific component in a combination or the concentration of a solution during scientific activity. The trick is to determine the color differences between various combinations and their absolute values. This is more instructive and scientifically valuable than relying on subjective judgments like the color of the solution. Colorimetry is used in a digital image-based (DIB) approach for determining active medicinal components. A computerized scanner with a controlled light intensity was connected to the detector. Different histograms were used to transform the photos. The colorimetric analysis of digital images provided for an easy-to-use and ecologically friendly method.


  • colorimeter
  • digital image based (DIB) colorimetric analysis
  • quantification of color
  • UV-vis spectrophotometer
  • reflectance spectrophotometer
  • green active pharmaceuticals

1. Introduction

When a light of occurrence with intensity (Io) passes from a solution, a portion of the light is revealed (Ir), a portion is absorbed (Ia), and the rest is transmitted (It),



Because of the measurement of (Io), (Ir) is omitted in colorimeters, and it is sufficient to regulate the (Ia). The light will be replicated (Ir) is kept consistent for this purpose by using cells with identical characteristics. The values of (Io) and (It) are then calculated.

The two fundamental equations of photometry, on which the colorimeter is based, show the mathematical link between the amount of light absorbed and the concentration of the substance.

Beer’s Law is a rule that states that if you drink.

The amount of light absorbed is directly proportional to the concentration of the solute in the solution, according to this rule.



as = Absorbency index.

C = Solution Concentration.


2. Lambert’s rule

According to Lambert’s law, the amount of light absorbed is proportional to the length and thickness of the solution under investigation [1, 2, 3, 4, 5].



A = Test Absorbance.

as = Standard absorbance.

b = the solution’s length/thickness.


3. Instrumentation

Components for Colorimeter,

The Colorimeter is a technique that measures the amount of absorbed light of a given wavelength in order to determine the concentration of a sample. Duboscq Colorimeter, created by Jules Doboscq in 1870, is one of the first and most popular designs. The components of a colorimeter are as follows:

  1. A source of light to illuminate the solution, commonly a blue, green, or red LED.

  2. Filters for light wavelengths of red, blue, and green.

  3. A slit through which a light beam can be focused.

  4. A condenser lens that divides a light beam into parallel rays.

  5. A cuvette in which the solution will be held. Glass, quartz, or plastic are used to construct.

  6. A photoelectric cell, which is a vacuum-filled cell that measures and converts transmitted light into electrical output. Light-sensitive materials, such as selenium, are used to create these.

  7. A transmittance or absorbance meter, either analogue (e.g., a galvanometer) or digital (e.g., a multimeter) [6].


4. Colorimeter’s process

  1. The LED or tungsten bulb emits white light to begin the procedure.

  2. A condenser lens concentrates this light into a parallel beam after passing through a slit.

  3. A series of lenses focus a certain wavelength of light on the solution as the beam passes through them.

  4. The problem answer is kept in a cuvette with a predetermined path length (width of the cuvette). The solution absorbs a portion of the light and transmits the remainder.

  5. The intensity of the transmitted light is transformed into an electrical signal, which is recorded by an analogue (e.g., a galvanometer) or digital meter [7].


5. The colorimetric principle

Colorimetry is the measuring of electromagnetic radiation’s wavelength and intensity in the visible spectrum. It’s widely used for determining the quantities of light-absorbing compounds and identifying them. The equation log can be used to represent the two laws when they are combined. I0/I = kcd, where I0 is the incident light intensity, I is the transmitted light intensity, c is the absorbing substance concentration, d is the distance through the absorbing solution, and k is a constant that varies depending on the absorbing substance, the wavelength of light used, and the units used to specify c and d.

The intensity of radiation passed through layers of various thicknesses of two solutions of the same absorbing substance, one with a known concentration and the other unknown, is a straightforward application of this term. The equipment is known as a photoelectric colorimeter when it compares intensities using a photoelectric cell rather than the eye.

The comprehensive noticeable band (white light) is frequently employed in colorimetry, The flattering color of the one captivated is seen as transmitted light. The instrument is called a spectrophotometer if it uses homochromatic light or a constrained band of radioactivity. It is used to demeanor dimensions in the ultraviolet and infrared regions as well as the visible spectrum. Colorimeters have mostly been superseded by spectrophotometers [8].


6. Color grading

Colorimetry is a term for the measuring of color. This field employs a wide range of instruments. The most advanced, spectrophotometers, measure the quantity of energy in each spectral wavelength. The emittance curves for light sources (see figure) and the reflectance curve for the paint color emerald green (see figure) are typical spectrophotometer results.

Electromagnetic radiation is defined as the movement of photons (also known as light quanta) over space, according to contemporary quantum theory. Photons are energy packets that travel at the universal speed of light at all times. The symbol h stands for Planck’s constant, and the value of corresponds to the frequency of an electromagnetic wave in classical theory. The numerical density of photons with the same energy h corresponds to the intensity of the radiation. The frequency of electromagnetic radiation influences these occurrences, as well as how it is made and viewed, how it occurs in nature, and how it is used in technology. The frequency spectrum of electromagnetic radioactivity varieties from extremely low values in the radio, television, and microwave ranges to noticeable light and elsewhere, too much sophisticated standards in ultraviolet light, X-rays, and gamma rays [9].


7. Color measurement aspects

A test bulb glows proceeding a bleached monitor, which is observed by an spectator, in a very ancient experiment. One or more of three lamps, each capable of producing light in three different colors, such as red, green, and blue, illuminate a neighboring section of the screen. The principal light is chosen at random, although it is tightly regulated. The observer can match the collective shade on the monitor to that of the test bulb by altering the intensities of these lights. The tristimulus value of a color is defined as the sum of the three primaries [10].

7.1 Color measurement is divided into three categories

  1. Visual matching and spectral matching are the two types of matching. Measurement in three colors. The information representative distinctive wavelengths collected from the noticeable bands of these things allows objects to be discriminated based on information representative distinguishing wavelengths achieved from the visible spectra of these substances, rather than tristimulus values. Spectrophotometer by immovable wavelength frequencies or specific shade devices are examples of the instruments used for this purpose.

  2. There are two types of colorimetric analysis: visual colorimetry and photoelectric colorimetry. There are two distinct methods to each method. The color absorption capacity is used to determine the amount of a colored component in a solution.

  3. To determine the true shade of the material in relations of tristimulus standards, which provides the shade a humanoid-like look.


8. Color matching

  1. Visual match: The simplest and oldest method of color measuring is visual color matching, which allows a shade to be projected but not restrained. It entails choosing or creating a shade that corresponds to the specimen’s X, Y, and Z tristimulus standards. The most straightforward method is to employ a large, regularly organized collection of color samples that can be visually matched by an observer alone or with the assistance of visual colorimeters, with the observer altering the reference color until it is judged to be equivalent to the specimen. This technique must be done in a well-lit environment. Standard color matching boxes with various illuminants can be found on the market and used to perform visual matching.

  2. Spectral matching: Spectral matching is the process of matching colors based on their wavelength The most precise approach for color measuring is spectral matching, which has two steps. The first, referred to as spectral photometry, entails measuring physical characteristics such as reflectance or transmittance curves, while the second, referred to as tristimulus assessment, entails recalculating or modifying the physical values derived from tristimulus data. Modern spectral measuring tools (spectrophotometers, spectrocolorimeters) can usually cover both stages in one run. By using this method, it is expected that 10 million hues may be distinguished. A monochromator, which is typically achieved in traditional structures using a grating, is at the heart of spectral color measurement devices. The slit width influences the wavelength bandwidth. Though the effect is the same, the most common method of covering the spectrum is to rotate the grating or prism rather than move the slit. The light sensitivity of the linear array detector, as well as optical fibers and concave gratings capable of focalizing light on the focusing plane where the array detector is situated, are used in today’s technology. This allows for the measurement of the entire light spectrum in a single shot while avoiding the use of spinning parts, and at a lower cost. Optical spectrophotometric scanners can now be used to do spectral matching under huge area samples. They’re spectrum imaging tools that can also be used to assess sample color consistency. The spectrum reflectance of the varnished layer is measured using spectral imaging, which is a physical quantity that is independent of the measurement apparatus. An ordered collection of the spectral reflectance of its pixels can be stored as a digital image based on this physical quantity. Spectral pictures have a number of benefits, including the following:

    1. Image reconstruction in CIE color space with illuminant selection and color matching functions

    2. Image replication with high fidelity using output devices such as monitors or printers that can work in more than trichromatic modes.

Colorimetry by spectrophotometer: The photoelectric method’s main premise involves converting the glowing energy conveyed by a stained system to rechargeable energy, which can be detected more easily, using a photoelectric detector. A light source with a constant intensity is required by the instrument.


9. Photometry of filters

  1. Color filters are used as a wavelength selection in this method, as indicated by the name. This strategy relies heavily on the selection of an appropriate filter. To increase sensitivity, the filter’s color should complement the color of the solution being tested. Most filter photometers produce a spectral bandwidth of 50 mj.i (or nm). (A micrometer is an insufficiently small unit.) Despite the fact that filter photometry provides more accurate and exact findings than the visual approach, it only covers a small portion of the spectrum compared to the spectrophotometer. As a result, shortened spectrophotometer is a term used to describe them.

  2. Spectrophotometry: A spectrophotometer measures a sample’s reflectance or transmittance as a function of wavelength. Which is a monochromatic that creates a spatially dispersed colored light spectrum on its focal plane. This allows you to choose any wavelength of incident radiant energy and determine the spectrophotometric curves of color samples radially. Interference from other color compounds can frequently be avoided by choosing the right wavelength.

  3. Trichromatic Colorimetry: According to researchers, the human eye has three receptors, and the differences in their responses contribute to color perceptions. Grassmann and Maxwell, on the other hand, were the first to assert unequivocally that color can be described mathematically in terms of three independent variables.

Color values and color difference measurements can be expressed in the following ways:

Newer benchtop instruments are also smaller, more automated, and less expensive than their forerunners. A new generation of entry-level benchtops has prices that are equivalent to portable computers. Software tools for color formulation and quality control have also made significant progress. Windows is used in the most, if not all, of the new debuts. They enable for customized display screen configurations and present electronically generated simulations of color standards and sample formulations in a variety of lighting conditions. Spreadsheet functionality, e-mail data transfer, and data transfer to and from portable spectrophotometers are among the other new features that make data manipulation easier [11].


10. Conclusion

Colorimetry has long been used in experimental, décor, pharmaceuticals, cosmetics, textiles, surface coatings, paper, ink, plastic, color photography, glass, and paint, as well as in biology as a health indicator and for cell and tissue examination. Color measurement is complicated by the fact that one can only ‘measure’ the reflection and transmission qualities of the materials that produce the color.

There are two methods that could be used: (i) color capacity of an purpose; (ii) strength quantity of reflected/communicated light at specific wavelengths The goal of this research was to create a tristimulus colorimeter with red, green, and blue leds, as well as to investigate their various uses. The architecture of the human visual system, color vision, and color vision theories are all covered in length in this chapter. We begin with an overview of color science and the internationally recommended CIE system, which serves as the foundation for colorimetric application, before moving on to color measurement instruments and color spaces used in color calculation and correction. This technique is used to quantify green active pharmaceuticals derived from natural sources.


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Written By

Vitthal V. Chopade and Jayashri V. Chopade

Submitted: 31 August 2021 Reviewed: 26 October 2021 Published: 20 July 2022