Liquid–liquid phase reaction between crystal violet and sodium hydroxide: kinetic study and precipitate analysis

To investigate reaction order and kinetic parameters of the reaction between crystal violet (CV) and sodium hydroxide (NaOH), various concentrations of the reactants were applied. The present work also verifies the unknown solid product produced under highly concentrated conditions. The reaction orders of CV and NaOH were determined to be 1 and 1.08 by pseudo rate method, respectively, with a rate constant, k, of 0.054 [(M−1.08) s−1]. In addition to pseudo rate method, the half-life approach was used to calculate the overall reaction order to verify the accuracy of pseudo rate method. The overall reaction order was determined to be 1.9 by the half-life method. The overall reaction order based on the two methods studied was approximately 2. The precipitate formation was observed when high concentrations of CV (0.01–0.1 M) and NaOH (1.0 M) were applied. Fourier transform infrared (FTIR) spectroscopy was used to compare the spectra of the precipitate generated and a commercial solvent violet 9 (SV9). Based on the FTIR spectra, it was confirmed that the molecular structure of the precipitate matched that of solvent violet 9.

SN, 0000-0001-9722-0170; YM, 0000-0001-5807-0544; TK, 0000-0002-0096-303X To investigate reaction order and kinetic parameters of the reaction between crystal violet (CV) and sodium hydroxide (NaOH), various concentrations of the reactants were applied. The present work also verifies the unknown solid product produced under highly concentrated conditions. The reaction orders of CV and NaOH were determined to be 1 and 1.08 by pseudo rate method, respectively, with a rate constant, k, of 0.054 [(M −1.08 ) s −1 ]. In addition to pseudo rate method, the halflife approach was used to calculate the overall reaction order to verify the accuracy of pseudo rate method. The overall reaction order was determined to be 1.9 by the half-life method. The overall reaction order based on the two methods studied was approximately 2. The precipitate formation was observed when high concentrations of CV (0.01-0.1 M) and NaOH (1.0 M) were applied. Fourier transform infrared (FTIR) spectroscopy was used to compare the spectra of the precipitate generated and a commercial solvent violet 9 (SV9). Based on the FTIR spectra, it was confirmed that the molecular structure of the precipitate matched that of solvent violet 9.
To collect the absorbance spectra, 50 µl of CV and 50 µl of NaOH solutions were pipetted into a microplate and placed into the UV-vis spectrometer. For the FTIR measurements, CV powder was mixed with a minimal amount of water to form a paste. If precipitates formed, the samples were separated from the solution and dried in an oven at 60°C for 48 h.

Characterization
The UV-vis spectra were obtained with a Tecan Infinite 200 PRO UV-visible spectrophotometer. The samples were pipetted (100-1000 µl Reference 2 micropipettes, Eppendorf ) into the microplate (96-well Corning Falcon 351172 STERILE R), and measurements were conducted at room temperature (21°C) in the range of 400-700 nm with a step size of 5 nm and two flashes. The general UV-vis spectroscopy measurement procedures are shown in scheme 1.
To quantify changes in CV concentration during the reaction, an absorbance band at approximately 590 nm was monitored until the peak intensity was indistinguishable from the baseline reference point of 650-700 nm [17]. All absorbance data during the reaction was recorded using the i-control™ Microplate Reader Software 1.11 (for the Infinite reader). In addition to UV-vis spectroscopy, to discern the time for the reaction to reach completion, an extra batch of samples was prepared, and the colour change was recorded. The infrared (IR) spectra were obtained using a Nicolet iS50 FTIR spectrometer (Thermo Fisher Scientific) equipped with an added attenuated total reflectance accessory. The FTIR spectra were recorded in the range of 400-1800 cm −1 at a resolution of 4 cm −1 with 32 scans. A background spectrum was collected before each sample was analysed. The spectra were obtained using OMNIC software.

Results and discussion
3.1. Effect of crystal violet and sodium hydroxide concentration on crystal violet decolorization and precipitate formation UV-vis spectroscopy was applied to analyse the CV concentration during the reaction with NaOH. As shown in figure 1, the maximum CV absorbance occurs at approximately 590 nm. As the concentration of NaOH was increased from 0.05 M to 0.5 M with a fixed concentration of CV (1.0 × 10 −4 M), the reaction time decreased from 40 min to 3 min 30 s (figure 1a-d). Note that when a CV concentration of 1.0 × 10 −4 M CV only was analysed (without NaOH), the absorbance peaks (500 nm-650 nm) showed extensive saturation, resulting in the absence of a clear peak (not shown for brevity).
The obtained results were used to calculate the kinetic parameters. For comparison purposes, the video captured images are shown in figure 1a 0 -d 0 ). As expected, the time needed for CV to decolorize decreased with increasing NaOH concentration. Figure 2 shows the UV-vis absorbance spectra with varied CV concentrations (1.0 × 10 −4 M and 1.0 × 10 −5 M), while the NaOH concentration was fixed at 0.1 M. As observed, the reaction time decreased from 20 min to 12 min 58 s with decreasing CV concentration. Although it was confirmed that decolorization efficiency can be improved with increasing NaOH concentration (figure 1a-d) or decreasing CV concentration (figure 2a,b), the reaction time was not fully dependent on the NaOH/ CV concentration ratio (figure 2c). In the case of the 1.0 × 10 −5 M CV and 0.1 M NaOH reaction (NaOH/CV ratio = 1.0 × 10 4 ), the reaction time for CV decolorization was longer than that of the 1.0 × 10 −4 M CV and 0.5 M NaOH reaction (NaOH/CV ratio = 0.5 × 10 4 ). This result suggests that the efficiency of the CV decolorization reaction could be controlled by modulating the NaOH/CV ratio and the absolute value of the NaOH (or CV) concentrations. In the case of the 1. royalsocietypublishing.org/journal/rsos R. Soc. Open Sci. 9: 220494 (NaOH /CV ratio = 0.1 × 10 4 ), based on the results shown in figures 1 and 2, it can be concluded that the absolute value of the NaOH concentration has more of an impact on the rate than the CV concentration.

Determination of reaction order and rate constant
CV is a relatively large molecule with three benzene rings and an amine bonded to a central carbon atom. Because NaOH is a strong base, it rapidly dissociates into Na + and OH − ions in solution, saturating the CV solution with hydroxide ions. A kinetic study of the decolorization of CV was performed based on the results presented in figures 1 and 2. The rate of the reaction is given by the generalized rate law: With an excess of NaOH (i.e. NaOH/CV concentration ratio = 1.0 × 10 3 -1.0 × 10 4 ), the rate equation could be simplified using the assumption that NaOH concentration can be considered constant throughout the reaction, ð3:2Þ The rate constant was determined using a method involving a pseudo rate constant (k 0 ) to simplify the rate law, The linear relationship between absorbance (A) and CV concentration is given by Beer's law, where ε is the molar absorption coefficient, c is the concentration and l is the optical path length.
To royalsocietypublishing.org/journal/rsos R. Soc. Open Sci. 9: 220494 time based on the integrated rate law, As shown in figure 3b-e, all datasets are linear when the natural log of [CV + ] is plotted against reaction time, indicating that the order dependence with respect to [CV + ] is 1st order, which is in accordance with previous studies [4,12,13,18].
While the conclusion of first-order kinetics with respect to [CV + ] coincides with previous literature findings, the precise values of k and n were calculated to further investigate the reaction kinetics. According to equation (3.6), the pseudo rate constant, k 0 , is determined by the slope of the linearized graphs found using the absorbance data. Obtaining pseudo rate constants for different concentrations of hydroxide allows a system of linear equations to be solved. Reaction order, n, with respect to [OH − ] and the reaction rate constant, k, are based on two solutions in this system. can be readily carried out to determine n and k. This approach improves accuracy by incorporating data from all three runs, rather than using only two runs, as reported in previous studies [15,16]. Therefore, when ln([k 0 ]) is plotted against ln([OH − ]), the slope will be linear. This slope provides the reaction order (n) with respect to [OH − ] and the y-intercept represents the k value.   figure 3g, the rate order with respect to [OH − ], n, was determined as 1.08 and the reaction rate constant, k, was determined as 0.054 (M −1.08 ) s −1 . This quantitative rate law allows the kinetics of the equation to be understood, as it is apparent from the reaction order that changing the amount of hydroxide affects the reaction rate more than changing the amount of CV would. These results are supported qualitatively as well, given that the hydroxide functional group is smaller than the CV molecule. More hydroxide in solution would lead to more frequent intermolecular collisions.
The obtained reaction order and rate constants were compared with previous studies, and the results are displayed in table 1.
Many previous studies determined that the reaction order in respect to both CV and NaOH is first order, which is well matched to the current results, while the obtained [OH − ] reaction order (1.08) was slightly higher than the reported value. The obtained rate constant, however, is quite different compared with the literature data, even at similar reaction temperatures. It is assumed that higher CV and NaOH concentrations under the current experimental conditions could contribute to the lower rate constant. For instance, most previous literature indicated that the concentration of NaOH fell between 0.0001 and 0.09 M [4,[13][14][15][16], while the current hydrolysis reaction was carried out at higher (0.1-0.5 M) NaOH concentrations. Highly concentrated solutions of NaOH may lead to a difference in the reaction rate constant, which requires further investigation. Based on the literature results, it is concluded that several reaction conditions such as mixing conditions, concentrations of reactants (CV or NaOH) and reaction temperatures, could contribute to the discrepancy of rate constants [4,[13][14][15][16]. The steady state or transient state condition may also affect the rate constant values. Thompson & Jason [14] reported that the first 30 s of data were ignored when calculating the reaction orders and rate constant, as the solutions were mixing in the cuvette during this time. Another variation in the rate constants may arise due to the selection of different CV peak wavelengths (i.e. 530, 590 and 595 nm) from the UV-vis spectra, although this effect is expected to be minor [13,14]. As shown in table 1, it is clear that the reaction rate is directly related to the reaction temperature. For instance, Salahudeen & Rasheed and Thompson & Jason studied the effects of temperature on CV decomposition and reported that the rate constant increased with increasing reaction temperature [4,14].

Overall reaction order from half-life method
Considering the error caused by the pseudo rate method, which assumes that the reaction order with respect to CV is 1, the half-life method was employed to calculate the overall reaction order of the hydrolysis of CV. This method assumes that the consumption of NaOH is proportional to the  royalsocietypublishing.org/journal/rsos R. Soc. Open Sci. 9: 220494 7 consumption of CV. Therefore, the consumption ratio (c) for the reactants is constant.
Thus, the hydrolysis reaction in equation (3.1) can be written as follows: wherek ¼ k Á c and x ¼ m þ n (overall reaction order). The following equation (3.11) represents the relationship between [CV þ ] and reaction time (t): Since the half-life of the reaction, t 1=2 , is defined as the time required for the reactant concentration to fall to half of its original value, equation (3.11) can be re-written as equation (3.12) and equation (3.13), Regression analysis on a plot of ln t 1=2 against ln [CV þ ] 0 can be readily performed to determine the overall reaction order x [19]. Figure 4a shows the CV conversion as a function of reaction time for different concentrations of NaOH. Considering the rapid reaction at the beginning of the hydrolysis, the sample with the lowest NaOH initial concentration, 0.05 M, was chosen for further analysis. As  royalsocietypublishing.org/journal/rsos R. Soc. Open Sci. 9: 220494 shown in figure 4b, the CV concentration decreased exponentially with increasing reaction time. Four points were chosen to calculate the overall reaction order (x). Figure 4c depicts the plot of ln t 1=2 and ln [CV þ ] 0 for the various CV initial concentrations and the half-life derived from figure 4b. Based on figure 4c, the overall reaction order was determined to be 1.90 by using the half-life approach. Although this value is slightly lower than that of the pseudo rate constant method, 2.17, the overall reaction order of the CV and NaOH reaction can be estimated to be approximately 2 at room temperature (21°C). This result is consistent with previous studies as shown in table 1.

Analysis of precipitate chemical
Although the reaction times varied with different CV and NaOH concentrations (figures 1 and 2), due to the excess of NaOH, most CV molecules were completely converted into a new compound, solvent violet 9 (SV9), as shown in figure 5 [18].
To confirm the existence of SV9, high concentrations of CV (i.e. 0.1 M and 0.01 M) and 1.0 M NaOH were applied. It should be noted that the precipitate had an intense dark colour ( photo in figure 6)), in contrast with the colourless solution ( figure 1a 0 -d 0 )) produced when the reactants were lower in concentration. This phenomenon is probably due to the non-spontaneous nature of the highconcentration reaction, as it did not proceed to completion with an excess of CV to decolorize. To further study the molecular structure of these precipitates, FTIR spectroscopy was used. For comparison purposes, solid CV and solid SV9 samples were analysed, with the results displayed in figure 6. The spectrum representing the untreated, solid CV contains strong peaks at approximately 1162 cm −1 , approximately 1349 cm −1 and approximately 1576 cm −1 , which correspond to C-N stretching vibration (or C-H stretching in aromatic ring), C-N stretching of aromatic tertiary amine (or C-H deformation in methyl group) and C=C stretching of the benzene ring, respectively [20][21][22][23]. The spectra of the precipitates show that most peak positions are very similar to those found in the   royalsocietypublishing.org/journal/rsos R. Soc. Open Sci. 9: 220494 spectrum for solid CV, while the intensity of the peaks changed drastically. For instance, the intensity of the approximately 1576 cm −1 peak in the CV spectrum decreased after the reaction, and the peak shifted to approximately 1564 cm −1 . Upon closer inspection, the precipitate spectra were found to contain a weak peak at approximately 1124 cm −1 corresponding to C-O stretching in the tertiary alcohol, which is not present in the CV structure [24]. Compared with the spectrum of SV9, the spectra of the precipitates clearly show that the peak positions and intensities are well matched. Therefore, it is reasonable to conclude that the product formed after CV reacted with NaOH is SV9.

Conclusion
In this work, the reaction between CV and NaOH was investigated using UV-visible spectroscopy, FTIR spectroscopy and video imaging. UV-vis spectroscopy was used for quantitative analysis, specifically for the reaction order and rate constant derivation, while video imaging was used for qualitative analysis. The reaction orders of CV and NaOH are 1.00 and 1.08, respectively, and the calculated rate constant (k) is 0.054 [(M −1.08 ) s −1 ]. Another method using the half-life approach determined the overall reaction order to be 1.9. Because the results differ only slightly, the overall reaction order of the CV and NaOH reaction can be estimated to approximately 2 at room temperature (21°C), which matches previous studies. FTIR spectroscopy was used to study the molecular structure and bonding vibration of CV, the precipitate, and SV9. When high concentrations of both NaOH and CV reacted, a precipitate formed, which was concluded to be SV9 by FTIR analysis.