TOPIC 13.

INDICATOR DYES

INTRODUCTION
Much work has been spent during the last 150 years to continously improve the stability of dyes. In
nature, however, not only color-stable dyes (e.g., indigo) exist but also dyes which change their color.
As early as 1660, Boyle observed that certain plant extracts changed their color on treatment with
acid or base. This effect is well known to gardeners in the case of hydrangea (or hortensia): the color
of its blossoms can be changed by watering it with slightly acidic or alkaline water. The first indicator
dye which came into use, was litmus , which was isolated from the orseille lichen, Roccella tinctoria.
It changes its color from red (acid) to blue (alkaline). The systematic investigation of natural colors led
to the discovery of several dyes and dye classes which are able to change their color. Thus, it was
found that the origin of the color of hydrangea blossoms is an anthocyan dye. Interestingly, in nature
these dyes are not always and necessarily used as indicators: the red color of roses and the blue
color of the blue cornflower are based on the same molecule (also an anthocyan), which in vitro
reacts as an indicator dye. In the plants however, this dye does not change color: the red rose does
not change to blue due to the presence of an intrinsic buffer system.

General Principles
In general terms, an indicator dye can be characterized as follows: (1) It shows different optical
properties when the system of which it forms a part changes its status, and (2) This change is
reversible. The first property means that the indicator molecule reacts as a part of the system. When
the number of [H3O]+ ions in this system is in a large excess compared to the number of indicator
molecules, the influence of indicator molecules can be neglected. When, however, the numbers of
[H3O]+ ions and indicator molecules become comparable, e.g., in very weakly buffered systems like
raindrops, the influence of the indicator molecules is significant. This phenomenon is known as
indicator error. The second property in particular distinguishes indicator dyes from colorforming
reagents. The latter are widely used in reactions in which colors are formed by chemical or
biochemical (mostly enzymatic) reactions between the analyte and added reagents. These reactions
are widely used in analytical and diagnostic test kits but are normally not reversible. The most
important structure changes for indicator dyes are (1) Reversible transition between acid and base
form of a molecule (pH indicator), (2) Reversible transition between reduced and oxidized forms of a
molecule (redox indicator), and (3) Reversible transition between the free molecule and its complex
with a cation (metal indicator).

CLASSES OF INDICATORS
pH Indicators
In common laboratory use, the term indicator is often employed to refer to pH indicators. A pH
indicator molecule exhibits an equilibrium between the indicator acid InA and the indicator base InB
[Eq. (1)].

To be suitable as an indicator molecule, the absorption spectra of InA and In B must be different. In
the transition from the acidic to the alkaline form (and vice versa), a point occurs where In A and InB
are present in comparable numbers. The consequence is a mixed color at this point. Important for the
practical application is that the complete transition requires a certain pH range. Therefore a pH
indicator does not change its color at a sharp pH value but within a range. Table 5.17 lists ranges and
colors of common pH indicators, and Figure 5.1. shows a graphical overview of the pH ranges and
color changes.
Individual pH Indicators. The most commonly used pH indicators include azo dyes, nitrophenols,
phthaleins, and sulfophthaleins. Varying the substituents on the chromophores yields pH indicators
with different transition ranges, and the complete pH range between 0 and 14 can thus be covered,
as shown in Table 5.17. Introducing acid groups (sulfo or carboxyl) leads to water-soluble indicator
dyes. pH indicators are generally used as 0.1% solutions in water, ethanol or water/ ethanol mixtures.
The special case of pH papers is described in Section 5.8.4; for pH indicators for nonaqueous
systems, see below. Structures of some of the most common pH indicators are shown in the
following:
Indicator Mixtu res (Multic omponent). Mixing several indicators of different transition ranges leads to
universal indicators which can be used over a wide pHrange. Table 5.20 lists examples of such
indicators.

Redox Indicators
Redox indicators are organic molecules whose oxidized and reduced forms show different colors.
The two levels of oxidation form a redox system which can be described by the standard potential or
rH value. A short definition of the two terms is given in [1]. Here, some more practical properties are
mentioned: . The redox potential of an indicator depends strongly on pH because hydrogen ions are
directly involved in the redox process, as the following example of Variamine Blue (7; C.I. 37255)
shows. . The color of redox indicators changes over a certain range. In addition, redox reactions often
proceed slowly. . The amount of redox indicator should be kept as small as possible because the
indicator itself is a redox system which might influence the system under investigation. This parallels
the indicator error of pH indicators mentioned above. The redox potential of the indicator system must
be chosen in such a way that the indicator does not react with the titrant: for an oxidizing titrant, the
redox potential of the indicator must be greater than that of the titrant. For a reducing titrant, the
reverse must be the case. Examples of redox indicators are Indigo Carmine, which is frequently
employed, and ferroin, an iron( II) complex with three 1,10- phenanthroline ligands (14), which is
widely used in water and wastewater analysis in the determination of chemical oxygen demand
(COD) in an internationally standardized procedure. For the structure of the organic ligand, see p.
539.

Metal Indicators

Metal indicators are organic molecules which form specifically colored soluble complexes with metal
ions in aqueous media. Here, the color of the complexes and of the free indicator must be different.
These reactions can be used in two analytical procedures: volumetry (complexometry) and
colorimetry/ photometry. In both methods, the concentration of metal ions is determined, but with
different techniques. Colorimetry/photometry exploits the fact that the color intensity of the complex
correlates to the metal ion concentration, at least within a certain concentration range. Therefore,
sufficient indicator is added to the sample that the indicator forms a colored complex with all ions of
the metal of interest in solution. The color intensity of this complex is then either compared visually
against the color of reference samples (colorimetry) or the color intensity is measured at a defined
wavelength with an instrument and then compared with a calibration curve (photometry). Volumetry
uses the formation of complexes between the metal ions and a complexing reagent (the titrant). This
reagent is added in the same manner as in titration. Here, the metal indicator serves as a classical
indicator: normally, a small amount of it is added before titration, and this leads to complex formation
with a certain number of metal ions, but not with all, in contrast to colorimetry/ photometry, and
consequently to a colored solution. Addition of the titrant then leads to formation of complexes
between this reagent and the metal ions which are colorless and more stable than the indicator.metal
complexes. As a result, the color of the solution changes when all free metal ions have reacted with
the titrant, because the next small addition of titrant forms a complex with the metal ions which were
so far in the metal.indicator complex. This complex is destroyed, loses its color, and the color of the
solution changes. The concentration of the metal ion is proportional to the added amount of titrant.
The main issue with metal indicators is selectivity. Generally, the selectivity can be improved by
optimizing the reaction conditions, that is, by addition of buffers, masking reagents, etc. Nevertheless,
many special reagents have been developed for very special applications in the last four decades,
mainly by users of photometric methods, who had no or only limited access to modern sophisticated
spectrometric methods. Chemical Structure. The complexes formed by the metal indicator and the
metal ions contain an electron donor and an electron acceptor part. If the donor molecule (ligand )
contains two or more atoms with donor properties, the complex is known as a chelate. Most organic
complexing agents can be regarded as bidentate ligands (they contain two donor atoms) and form
five- or six-membered chelate rings. The electron-donating atoms in these chelates include oxygen,
nitrogen, and sulfur. The other part of the molecule varies significantly, as the examples below show.
The two following examples demonstrate how the general structure of the molecule determines its
properties:

Both molecules contain the same donor atom configuration. Diacetyldioxime is highly specific for
nickel and palladium. If the same donor atom configuration is incorporated into a heterocyclic system,
as in the phenanthrolines, the ligands become specific for copper and iron. This specificity can be
directed by choice of the substituents R in the molecule: if R = H, the ligand is specific for iron; with R
= CH3 and/or C 6H5, the ligand is copper-specific. Some of these metal indicators are used as
reagents in photometric or visual test kits for the detection of metals. Here, these molecules act like
color-forming reagents but are still indicators.

Indicator Papers
These papers are obtained when uncoated unfilled paper is soaked with solutions of pH indicators
and then carefully dried. Depending on the end use, one or more indicator dyes can be applied.
Three types of indicator papers are commercially available. Simple indicator papers only show if a
solution is acidic, neutral, or alkaline. Examples of this type are litmus paper, phenolphthalein paper,
and congo paper. Universal indicator papers permit pH determinations over the total pH range from 0
to 14. The determination of the pH is made by comparing the color of the paper with a reference color
scale. The accuracy is ca. 1 pH unit. Special indicator papers cover ranges from 2 to 5 pH units. The
determination method is the same as with universal indicator papers. The accuracy is better: 0.5 units
is regularly attained; under favorable conditions, 0.2.0.3 units is possible.
Nonbleeding Indicator Papers
It can be prepared either by using appropriate direct dyes or reactive dyes. In the latter case, pure
linters (the raw material for high-quality paper) are suspended in water, and the solution of the
reactive dye is added. For example, the dye 15 reacts via the sulfonic acid group in the side chain.
The reaction mixture is then made alkaline and the dye reacts with the linters. After completion of the
reaction, the fiber pulp is centrifuged, washed electrolyte-free, and processed on a paper machine to
form the pH paper. Paper produced in this way is mostly bonded onto a plastic material and used as
pH test strip.

pH papers and strips with covalently bonded indicator dyes can remain almost indefinitely in solution
without any migration or dissolution. Thus, pH determinations in hot water (up to ca. 70 °C), alkaline
solutions, and weakly buffered solutions become possible. Also, pH measurements in turbid solutions
or suspensions are possible, as the paper can be briefly rinsed with distilled water without changing
the color. Experiments have been performed with other carriers such as hydrophilic plastic foils and
even textiles, but none of these products has come to market.