Comparison of efficiency of natural and artificial indicators

I always have an inclination for performing an experiment and collect data as that would really allow me to grow my self-management skill. So, the challenge was to arrive at a suitable topic for this. During the times of COVID, I have seen various ayurvedic (an ancient therapeutic science in India) and naturopathic treatments to build up immunity. Including that in my own family, I have seen people pondering over the fact that whether to go for the allopathic medicines or choose the natural alternatives. Incidentally, at the same time, I was introduced to the concept of – ‘Green Chemistry’ which in simple words speaks of greener and natural substitutes of synthetic chemicals. The concept that immediately clicked into my mind is to compare and contrast the effectiveness of natural substances over synthetic alternatives. But which category of substances should I choose? The choice was difficult as it has to be something which is easily available and the analytical procedure to compare the efficiency must also be not very complicated. Retracing back to my IGCSE Chemistry classes, I can remember of the natural indicators that can be used as an alternative for the synthetic indicators like methyl orange, phenolphthalein and methyl orange which are mostly used. Analysis of materials and quantification is an important aspect of chemistry. Continuous researches have offered more than one alternative for the determination of a particular parameter. For example, to determine the acidity of ocean there are so many methods like – using pH sensors, measuring levels of carbon-dioxide and so on. This inspired me to understand and evaluate how the choice of materials involved in an analytical procedure may impact the result obtained from it. Thus, I thought of conducting the titration between the same acid and base using various indicators to see which one would be more reliable and how the reliability can be ascertained. Thus, I arrived at the research question stated below.

How does the efficiency of natural indicators (turmeric, tomatoes, beetroot, cherries and Chinarose) measured in terms of pH range and pH at equivalence point differ from that of a synthetic acid base indicator-phenolphthalein, determined using a pH curve?

An acid base indicator is an organic acid or base where the conjugate acid base pair differ in colors. For example, if HX is taken as a weak acid that can acts as an acid base indicator and MOH as a weak base that acts as an acid base indicator, the equation of dissociation can be presented as:

HX (aq) + H₂O (l)\(\leftarrow\)--\(\rightarrow\) H₃O⁺ (aq) + X^- (aq) ................ (equation -1)

B (aq) + H₂O (l)\(\leftarrow\)--\(\rightarrow\) BH⁺ (aq) + OH^- (aq) .................. (equation-2)

The color displayed by the conjugate acid and base differs. Thus, the color of the solution actually depends which one of the conjugate form–the acid or the conjugate base predominates. The change of pH of the medium dictates the position of the equilibrium and thus the color of the solution as well.

According to the equation (1)

Acid dissociation constant : k_a = \(\frac{[H^+][X^-]}{[HX]}\)

At half-equivalence point, the concentrations of the conjugate base and the undissociated acid is same. Therefore, [X-] = [HX]

k_a = [H⁺]

Taking negative logarithm on both sides,

− log k_a = − log[H⁺]

pk_a = pH

The calculation above shows that at a pH = pKa of the indicator, the indicator works the best.

k_a = \(\frac{[H^+][X^-]}{[HX]}\)

H⁺ = \(\frac{k_a\ ×\ [HX]}{[X^-]}\)

− log[H⁺] = − log k_a + log \(\frac{[X^-]}{[HX]}\)

pH = pk_a + log\(\frac{[X^-]}{[HX]}\)

The color change of an indicator is prominent under two conditions

The concentration of the conjugate base is 10 times the concentration of the weak acid.

[X^-] = 10 [HX]

pH = pk_a + log\(\frac{[X^-]}{[HX]}\)= pH = pk_a + log\(\frac{[10HX]}{[HX]}\)= pk_a + log 10 = pk_a + 1

The concentration of the weak acid is 10 times the concentration of the conjugate base

[HX] = 10 [X^-]

pH = pk_a + log\(\frac{[X^-]}{[HX]}\) = pH = pk_a+ log\(\frac{[X^-]}{[10X^-]}\) = pk_a + log\(\frac{1}{10}\) = pk_a − 1

Thus, the pH range of an indicator is pk_a + 1 to pk_a - 1. It means that for an indicator to be suitably used in an acid base titration, the pH at the equivalence point must lie within ±1 of the pK_a value of that weak acid used as an indicator and pK_b for a weak base used as an indicator. For example, for methyl orange, a weak acid used as an indicator, the pk_a is 3.46. Thus, it is appropriate for an acid-base titration where the pH at the equivalence point lies within (3.46 + 1.00)= 4.46 to (3.46 – 1.00) = 2.46

The way, the weak acid HX changes it’s color depending on the presence of excess H+ ions in an acidic medium or excess of OH- ions in the basic medium can be explained using the Le-Chateleir’s principle and the equilibrium shown below

HX (aq) + H₂O(l)\(\leftarrow\)- \(\rightarrow\) H₃O⁺ (aq) + X^- (aq) ..........equation-3 

In an acidic medium, there is excess of Hydronium ions (H₃O⁺). This causes the equilibrium in equation-3 to shift more towards the reactant and thus the color of the acid (HX) predominates. Thus, indicator HX will show Color-A in acidic medium.

In a basic medium, there is excess of hydroxyl ion in the medium. This combines with the H₃O⁺ which is liberated from the dissociation of the weak acid HX and form H₂O as shown in equation-4. As a result, the equilibrium concentration of H₃O⁺ decreases.

OH- (aq) + H₃O⁺ (l) ------\(\rightarrow\)2 H₂O (l) ..........equation-4.

This reduces the concentration on the product side and thus the equilibrium shifts towards the product according to the Le-Chateleir’s principle. Thus, the conjugate base X- will predominate and Color-B will be shown.

A pH curve is obtained during an acid base titration where the pH of the analyte is measured as a function of the volume of the titrant added. It is presented graphically in the form of a scattered plot with pH along the y axes and the volume of titrant added along the x axes. A pH curve can be used to procure a lot of analytical information like volume of titrant required at the equivalence point to calculate the concentration of the analyte, pk_a of the acid / base used as an analyte or titrant provided it is weak. In a pH curve, an inflexion point is obtained around the equivalence point when the pH changes sharply on addition of a drop of the titrant. The mid-point of this region where the pH changes rapidly is marked as the equivalence point. A perpendicular can be drawn from the equivalence point to the y axes to deduce the pH at equivalence point and a perpendicular can be drawn from the equivalence point to the x axes to deduce the volume of titrant required at the equivalence point.

A pH curve can be mathematically treated to obtain the equivalence point. The equivalence point is basically the inflexion point of the curve. Thus, the polynomial equation that the pH curve follows can be used for a double derivative and the value of y axes (pH) and x axes (volume of titrant added; NaOH in this case) can be deduced.

Depending on the type of the acid and base used, there are four different types of pH curves

To delineate a comparative study of the indicators used, a pH curve will be used for the titration of a strong base NaOH with a strong acid HCl. The same titration will be performed using various indicators. From the pH curve, the following will be computed for each case, the pH at equivalence point, the volume of NaOH required at equivalence point, the pH range and also the color change. This method is known as pH metric titration.

This investigation is focused on four natural indicators- turmeric, tomatoes, cherries and beetroot. All of them contain some organic chromophores. These are organic molecules having extended conjugation (alternate single and double bonds) which enables them to absorb radiation in the visible spectrum and thus display the complementary color. Presence of Hydrogen ions or hydroxide ions causes them to alter their structural features and thus the color displayed also changes. The pigments present in various fruits are-

The natural indicators are equally efficient as the synthetic indicator – phenolphthalein and there is no significant difference in the efficiency of the different varieties of the natural indicator- turmeric, cherries, tomatoes and beetroot.

The synthetic indicator phenolphthalein is more efficient than the natural indicator. The natural indicators differ in their efficiency.

• A safety masks was worn so that the acid vapors are not inhaled as they may cause nausea.
• A laboratory coat was used to prevent exposing any of the chemicals to skin.
• Hair was kept tied so that it does not spill over the solutions made and used.
• The knife was used very carefully while cutting the fruits.
• The mixer blender was operated with care so that the paste does not spill out.
• Sanitizers were used as a part of the COVID protocol and physical distancing protocols were also maintained.

As the investigation involves the use of eatable substances, it was ensured that there is minimum wastage and to do so, the pulp left as a residue after filtering was used instead of disposing them. All COVID protocols were maintained while performing the investigation.

The procedure is such that it does not involve the emission of any green-house gas or major toxic wastage.

• The turmeric was cut into small pieces using a knife on a cutting board.
• The cut pieces of turmeric were inserted into the mixer blender and blended to obtain fine thick paste.
• The paste was transferred from the mixer grinder to a 100 cm³ glass beaker and distilled water was added to it.
• A glass rod was used to stir and make a suspension.
• The supernatant of the suspension thus obtained was filtered using a funnel and the filter paper. The filtrate was collected and used as the indicator solution.

The other indicator solutions were prepared in the same way. For Chinarose, petals of the flower was used.

• Take a watch glass and weigh 0.40 ± 0.01 g of NaOH pellets in it.
• The weighed solid was transferred to a 100 cm³ glass beaker.
• Distilled water was added till the mark of 100 cm³.
• A glass rod was used to stir the solution.

• A 1.00 graduated pipette was used to transfer 0.90 ± 0.05 cm³ of concentrated HCl to a 100 cm³ glass beaker which is already filled in with distilled water.
• Distilled water was added till the mark of 100 cm³ in the same beaker.

• A burette was taken and filled with 0.10 mol dm^-3 NaOH solution till the mark of 0.00 cm³.
• A 100 cm³ glass beaker was taken and 20.00 ± 0.05 cm³ of 0.10 mol dm^-3 HCl was added to it.
• 2.00 ± 0.05 cm³ of the phenolphthalein solution was added using a graduated 10.00 cm³ glass pipette.
• The pH probe was calibrated using the standard buffer solution of pH 4.00.
• The pH probe was inserted in to the glass beaker.
• 2.00 ± 0.05 cm³ of NaOH was added from the burette and the reading in the pH probe was noted down.
• Step-5 was continued until entire NaOH solution was added and after addition of 2.00 ± 0.05 cm³ of NaOH, the reading in the pH probe was recorded.
• The same process was repeated for two more times.
• The same steps were executed for other indicators – turmeric, tomatoes, cherries, Chinarose and Beetroot.

Refer to Appendix for other raw data tables – Figure - 21 to Figure - 25.

Sample Calculation when volume of NaOH solution added would be 0 cm³

Mean pH = \(\frac{4.52\ +\ 4.55\ +\ 4.53}{3}\) = 4.53

Standard Deviation = \(\sqrt\frac{{(4.53\ -\ 4.52)^2\ +\ (4.53\ -\ 4.55)^2\ +\ (4.53\ -\ 4.53)^2}}{3}\) = 0.01

Percentage Uncertainty = \(\frac{±0.01}{4.53}\) × 100 = 0.22%

y = −0.0003x³ + 0.0225x² − 0.2482Ux + 5.0833

\(\frac{dy}{dx}\) = −3 × 0.0003x² + 2 × 0.0225x − 0.2482

\(\frac{dy}{dx}\) = −0.0009x² + 0.045x − 0.2482

\(\frac{d^2y}{dx^2}\) = −2 × 0.0009x + 0.045

\(\frac{d^2y}{dx^2}\) = −0.0018x + 0.045

To determine the inflection point:

\(\frac{d^2y}{dx^2}\) = 0

−0.0018x + 0.045 = 0

x = 25.0 cm³

To determine pH equivalence:

y = −0.0003x³ + 0.0225x² − 0.2482x + 5.0833

y = −0.0003(25.0)³ + 0.0225(25.0)² − 0.2482(25.0) + 5.0833

y = 8.25

For turmeric,

pH at equivalence point = 6.75 ± 0.01

Percentage error in pH at equivalence point = \(\frac{±\ 0.01}{6.75}\) × 100 = 0.15

Volume of NaOH required at equivalence point = 24.00 ± 0.05 cm³

Percentage error in volume at equivalence point = \(\frac{0.05}{24.00}\) × 100 = 0.20

How does the efficiency of natural indicators (turmeric, tomatoes, beetroot, cherries and Chinarose) measured in terms of pH range and pH at equivalence point differ from that of a synthetic acid base indicator-phenolphthalein, determined using a pH curve?

The investigation will compare the action of the indicators based on three facts:

Volume of NaOH required at equivalence point

Concentration of HCl used = 0.10 moldm^-3

Volume of HCl taken = 25.00 cm³

Volume of NaOH required = \(\frac{100\ ×\frac{25}{1000}}{0.10}\) = 25.00cm³

Ideally, the volume of NaOH required at equivalence point must be 25.00 cm³ . As reported in Figure - 21, this expected value is obtained while phenolphthalein has been used as an indicator. For none of the natural indicators, this value has been obtained. Among all the natural indicators, turmeric and tomato gives a value of 24.00 cm³ which is closest to the ideal value. Maximum deviation has been found for cherry which gives a value of 22.60 cm³ . For the other two – beetroot and Chinarose, the values are 22.60 cm³ and 23.00 cm³ . Thus, if we consider that closer the value of volume of NaOH at equivalence point to the ideal value which is 25.00 cm³ , more efficient the indicator; the natural indicators arranged in descending order of efficiency should be turmeric ~ tomatoes, Chinarose, Beetroot and cherries.

pH at equivalence point

NaOH reacts with HCl according to the equation stated below

NaOH (aq) + HCl (aq) -------\(\rightarrow\) NaCl (aq) + H₂O

As the salt formed is NaCl which is a neutral salt, ideally the pH at equivalence point should be 7.00. Here, the pH at equivalence point is found to be in the basic region for phenolphthalein, Beetroot and Chinarose (values are 8.25, 8.77 and 8.99 respectively). While for the indicators, turmeric, tomato and cherry, the values are 6.75, 6.76 and 6.29 respectively. Thus again, turmeric and tomatoes can be claimed to be a better indicator than the others as they display a pH value closer to the ideal value which is not obtained in other cases. However, based on the justification that the values obtained for phenolphthalein, beetroot and Chinarose are way more than the ideal value 7.00, these indicators are definitely not claimed to be a suitable choice in this particular titration.

Color change at equivalence point

To discuss the color change, we need to refer to Figure - 9 of qualitative data. As indicated in that, all the indicators are showing different colors before and after the equivalence point and can thus be claimed to be suitable or appropriate for this titration. However, for some indicators multiple changes of color has been observed which may create a confusion. For example, in case of beetroot the color initially changes from magenta to fuchsia to rosewater to lemonade to colorless. If there is so many transitions of color, detection of end point will become difficult. Thus, in that way using this indicator will not be suitable for this titration. However, using a pH curve can resolve this issue as there the equivalence point is determined using mathematics from the pH curve. This issues is also there in using tomato and turmeric as an indicator.

As it is clear from the discussion in the data analysis section above, the efficiency of the natural indicators and that of the synthetic indicator is not same. Moreover, in respect to various factors the efficiency of all the natural indicators is also not the same. Thus, the null hypotheses is rejected and the alternate hypotheses has been accepted.

Acid base indicators are weak organic acids or bases. However, all the natural indicators used here are not the same. Curcumin has two phenolic-OH groups which can act as mild acid while Lycopene- the pigment in tomato is a hydrocarbon. Betaine, the pigment in beetroot has carboxylic acid groups which can lose a proton and behave as an acid. While the pigments in Chinarose and cherries are both polyscacchraides units and do not behave like a weak organic acid or base. Thus, the mechanism in which these pigments would show variation in color due to the presence of excess H+ or OH- in the medium would not be simply due to the shift in the position of equilibrium of their dissociation into a conjugate base or acid. This can rather be explained in the light of the gap between the HOMO (Highest Occupied Molecular Orbital) and (Lowest Occupied Molecular Orbital) which may have reduced or increased due to variation in the acidity or basicity of the medium. As the gap between the HOMO and LUMO increases, the energy absorbed will be of a greater frequency and thus the wave emitted as a complementary color will be longer.

• The values of volume of NaOH added have been varied at an interval of 2.00 cm³ . This gives the chance to have uniformly distributed and large number of data points in the pH curve. As the number of data points in a pH curve increases or is sufficiently high, the results interpreted from it becomes way more reliable and accurate.
• The equivalence point can be determined from a pH curve manually by estimating the mid-point of the part of the line where a jump is seen. However, here the equation of trend line has been obtained using MS-Excel and the double derivative method has been used to identify the inflexion point or the equivalence point. This makes the result more mathematically valid and acceptable.
• All significant factors like volume of indicator added, type of acid and base used in the titration has been significantly controlled so that it does not affect the accuracy of the data collected.

• There is no standardized protocol to measure the efficiency of an indicator. Thus, the analysis of this investigation is mostly qualitative in nature and does not have any quantitative aspect based on which a definite answer to the research question can be deduced.
• The investigation intends to understand the action of the pigments or organic dyes present within these natural substances as an acid base indicator. However, the indicator solutions used are simple aqueous extract and thus they surely contain chemicals other than the main ingredient or the dye under study. This may interfere with the accuracy of the result and is thus a major methodological limitation.

As efficiency of an indicator is a factor to assess the accuracy of data collected from a pH metric titration, there are many other factors that describes the accuracy of the result of an acid-base titration. For example, the analytical methodology followed in a titration between an acid and a base also plays a role. Titrations can be conducted in various ways like pH titrations, conductometric titration (by measuring electrical conductivity of the analyte against the volume of titrant added), potentiometric titration (by measuring the voltage of the analyte against volume of the titrant added), thermometric titration (by measuring the temperature of the analyte against the volume of base added). A specific pair of acid and base – ethanoic acid and NaOH for example can be taken where the concentrations of both the analyte and the titrant is known. Using the same volume of analyte (ethanoic acid) it can be titrated against NaOH solution in different ways- conductometric titration, potentiometric titration, thermometric titration and pH metric titration. In all cases a scatter graph with volume of base along the x axes can be obtained. From the inflexion point (for pH curve) or intersection point of two straight lines (as in case of conductometric, potentiometric and thermometric titration), the volume of titrant at equivalence point can be calculated. Using that, the concentration of the analyte can also be calculated. Thus, the percentage accuracy of the method can be calculated using the formula

Percentage accuracy = \(\frac{Calculated\ concentration\ of\ analyte\ -\ Actual\ concentration\ of\ analyte}{actual\ concentration\ of\ analyte}\) × 100

This will allow to address the research question- “ Does the percentage accuracy of the concentration of the analyte determined in a titration depends on the titration method-( pH metric, conductometric, potentiometric and thermometric ) used?”

Chemical Composition Functional Properties and Processing of Beetroot.

https://www.ijser.org/paper/Chemical-composition-functional-properties-and-processing-of-%20Beetroot.html.%20Accessed%2018%20May%202021.

Majolo, Fernanda, et al. “Approaches for the Treatment of Neurodegenerative Diseases Related to Natural Products.” Studies in Natural Products Chemistry, vol. 69, Elsevier, 2021, pp. 1–63. DOI.org (Crossref), doi:10.1016/B978-0-12-819487-4.00014-8.

University of Ljubljana, Faculty of Natural Sciences and Engineering, Department of Textiles, Graphic Arts and Design, Snežniška ulica 5, SI-1000 Ljubljana, et al. “The Influence of Mordanting on the Dyeability of Cotton Dyed with Turmeric Extract.” Tekstilec, vol. 58, no. 3, Sept. 2015, pp. 199– 208. DOI.org (Crossref), doi:10.14502/Tekstilec2015.58.199-208.

Vankar, Padma S., and Dhara Shukla. “Natural Dyeing with Anthocyanins from Hibiscus Rosa Sinensis Flowers.” Journal of Applied Polymer Science, vol. 122, no. 5, Dec. 2011, pp. 3361–68. DOI.org (Crossref), doi:10.1002/app.34415.

Yan, Zhiying, et al. “Visible-Light Degradation of Dyes and Phenols over Mesoporous Titania Prepared by Using Anthocyanin from Red Radish as Template.” International Journal of Photoenergy, vol. 2014, 2014, pp. 1–10. DOI.org (Crossref), doi:10.1155/2014/968298.