QUANTITATIVE DETERMINATION OF COPPER (II) CONCENTRATION BY SPECTROPHOTOMETRY

R. Lee1, I. Fernando1, H. Dumo1, R. Villacorta1 and M. Embalsado1

1INSTITUTE

OF BIOLOGY, COLLEGE OF SCIENCE UNIVERSITY OF THE PHILIPPINES, DILIMAN, QUEZON CITY 1101, PHILIPPINES DATE SUBMITTED: MARCH 15, 2013 DATE PERFORMED: MARCH 8, 2013

ABSTRACT Spectrophotometry is a photometry that deals with the visible and ultraviolet light. It is a chemical analysis used to determine the absorbance or concentration of a solution. It is governed by the indirect relationship of the transmittance of a monochromatic light and its absorbance as its passes through an aqueous solution of a certain concentration (Brown, 564). In the experiment, the Beers Law was used to ascertain the copper (II) of a sample. The instrument used is the spectrophotometer, which determined the absorbance of a solution from its light transmission. Plotting the absorbance versus concentration data made a calibration curve that was said to be in a linear form, which holds true for the Beers laws concepts. A working equation was derived and the samples Cu 2+ concentration is revealed to 2574 ppm, which has 2.94 % deviation from the hypothetical ppm of 2500. The RSD is zero and the confidence interval is 2574 ppm. The very accurate and precise results prove the efficiency of spectrophotometry in chemical analysis in ideal working conditions and apparatuses.

INRODUCTION In the early 17th century until the industrial revolution, the pursuit of chemical analysis was a strong interest for researchers and scientist all over the world. It has then progressed exponentially and branched out to entirely different methods of chemical scrutiny. A good example of this radical improvement is the application of colorimetry, the science to describe physically the human color perception, to the function of molecular absorption spectroscopy. Nessler tube, a precolorimeter instrument, was developed in early 1830s. This instrument determines the concentration of an analyte by comparing the samples color to a set of standards using the tubes. Although the development of analysis with the use of the nessler tubes is a huge leap for separation science, it is still limited by the tedious nature of visual color comparisons. In the 1930s and 1940s, advancement in electronics and application of colorimetry brought new light to spectrophotometry; a science that deals with the ultraviolet and visible light transmittance and absorbance of an analyte as a function of its concentration. The modern instruments for spectrophometry called spectrophotometers now use monochromators for wavelength selection, which

in turn will increase the accuracy of the analysis. (Harvey, 376) This was developed and invented by Arnold J. Beckman in the National Technologies Laboratory (NTL) and started by the company Beckman, this would soon be the legacy of the said company and NTL. The concepts defining the use of the ultraviolet and visible light spectrophotometer, shortened as UV-Vis spectrophometer, are all dependent on the properties of light and its absorption. The whole experiment was summarized to the notion that: molecules and atoms absorb and refract light, this would be fundamental structure of spectrophotometry and its concepts. When light passes by or hits an object, some of it are refracted and others are absorbed, for an instance, in a copper solution, the Cu2+ ions reflects the blue spectrum and absorbs the other colors. This would also mean that, the more copper ions present in the solution, the higher is the absorption of light in the solution. In spectrophotometry, the absorption would be a function of light transmittance as it passes through an aqueous solution.

Figure 1. Light Transmittance In figure 1, light was transmitted through a cell containing an aqueous solution, as you can notice, the figure emphasizes the thinning of the arrow line after it passes through, this is because absorption of light took place in the solution. PO is the source of radiance or light and PT is the transmitted light, which is now attenuated or decreased in degree of energy. The weakening of the intensity of light is quantified by two terms, namely: transmittance and absorbance, which are related indirectly in proportion. Transmittance, defined as the ratio of the traversing light and transmitted light (1), is commonly expressed as a percentage called transmittance percentage (2). It is simply the degree of transformation of the initial radiation to transmitted light.

the activity of the ions and molecules of the solution, its just the limit of how the valence electrons of the ion could move from higher orbitals or energy levels. Concentration, as discussed before, follows the logic of absorbance and quantity: the more ions, the absorption would take place. Lastly is the path of length, it would be explained by probabilities, as light passes through a longer path on the solution, the probability of interacting with the molecules and its absorption would be higher (Skoog, 718-720). Assimilating all these relationships and proportionality, we would now derive the Beer-Lambert-Bouguer Law or simply Beers Law:

Named after scientists August Beer, Johann Heinrich Lambert and Pierre Bouguer, the Beers Law (5) relates absorbance of the solution directly to the three factors mentioned before, the absorptivity which is denoted as a or the greek letter when the concentration is in molar units, the path length which is b, concentration which is represented by c. This expression and the concepts discussed before would be utilized by the Uv-vis spectrophotometer, a singe beam of light would pass through cells with different known concentrations of Cu2+ to determine each solutions absorbance, the data would then be regressed linearly to determine a working equation. To further understand the mechanics of the instrument, take a close look at Figure 2.

Absorbance on the other hand, is the quantity of absorbed energy of the solutions molecules and ions on the passing beam of light. As light passes through an aqueous solution, electrons of the atoms and ions present are excited and placed to higher energy states; this is due to absorbance itself. In visual representation, the absorbed energy would be the colors that do not manifest to our vision ass we look at it, and transmitted light are the colors that we see. We could deduce that the higher the absorption, the lower or weaker is the transmittance, this relation could be expressed with equation 3 and 4, where A is the absorbance.

Figure 2. Spectrophometer Mechanics The main objective of this experiment is to find the concentration of a Cu2+ solution and then find the volume of the initial analyte, to determine this; the determined working equation and the absorbance of the copper solution with unknown molarity was used. The same cell was used to keep the path length constant and since the sample is at the same composition and working conditions, its absorptivity barely changed. Knowing these facts, the absorbance determined by the spectrophometer was injected to the working equation between the relationship of the

The absorbance and transmittance of light are mainly affected by three factors; they are the analytes concentration, the absorptivity of the components and the path length of light as it traverses the solution. The absorptivity mainly is just a constant that depends on

absorbance and concentration, multiplying it with the dilution factor would then give us the sought after molarity of the sample analyte. METHODOLOGY The concentration of unknown sample was analyzed through the determination of the relationship of absorbance and concentration. Seven 50 mL solutions are prepared from a 250mL 2500ppm Cu2+ and concentrated NH3 solutions. Table 1 shows the composition of each working solution. Table 1. Absorbing Analytes Added 2500 Added ppm Cu2+ concentrated NH3 solution (mL) (mL) Solution 1 0.00 10.00 Solution 2 2.00 10.00 Solution 3 4.00 10.00 Solution 4 6.00 10.00 Solution 5 8.00 10.00 Solution 6 10.00 10.00 Sample 7.4 10.00 Solution The said components are added to a 50 mL volumetric flask. Copper (II) is the absorbing particle and NH3 is the color-intensifying particle. The reason for the addition of ammonia is to intensify the color of the solution; this will amplify the absorbance of the solution and will yield to better results. After the addition, the solutions are diluted to mark and then covered with parafilm. The solutions were then subjected to the Uv-Vis spectrophotometer. The analytical wavelength or the wavelength where the absorbance of the solution is at its peak was first determined. The most concentrated solution was used to maximize the absorbing capacity of the solution, which is solution 6 for this experiment. Solution 6 was poured in the cell and the instrument was turned to spectrum mode. The wavelength range was set to 300nm to 700nm, the visible light range. The instrument scanned and the analytical wavelength was determined, this would be used for the absorbance reading of the other solutions since this is where absorbance is maximized and error possibility is at its lowest. The next step was to eliminate the effect of ammonia to the absorbance of the solution, in order to do this, solution 1 or the blank solution in the cuvette was scanned and the value was put to autozero.

The absorbance of solutions 1-6 was then determined. The order is from the lowest to the highest copper (II) content to minimize concentration increase for each analyte. Lastly is the determination of the samples absorbance, which was done in triplicate. RESULTS AND DISCUSSION The analytical wavelength is 596 nm. This means the solution is the hue of blue-green. This is the monochromatic light that was used in the experiment (Jeffrey, 660). In the introduction, the concepts of spectrophotometry were discussed; it was then used to collect the needed data for the graph of the best-fit line and its equation. 0.6 A 0.5 b 0.4 s o 0.3 c r 0.2 e b 0.1 a 0 n

y = 0.001x - 0.005 R = 1 0 200 400 Concentration 600

Figure 3. Relationship of absorbance of Cu2+ solutions to their corresponding concentrations As conferred before, the absorbance of an analyte is proportional to its concentration. Absorbance values from the copper solutions with known concentrations are collected and graphed vs. each parts per million or ppm of a solution, a trend line and a corresponding equation were determined. The equation was then used to determine the concentrations of copper solutions with known absorbance, x would be the value of concentration and y would be the absorbance. The equation for the relationship of the absorbance and concentration of the Cu2+ solution is derived to be:

Concentration used absorbance instead of transmittance in quantification due to convenience with the linear relationship between A and c. This is the reason why the Beers Law is expressed in terms of absorbance rather than the transmittance. Since there is already a working equation, the concentration

of the working sample solution and the initial analyte solution was calculated. Trial Table 2. Sample Analysis Absorbance Cu2+ ppm in Working Sample 0.386 381 ppm 0.386 0.386 0.386 0 0.386 Volume Percent error 381 ppm 381 ppm 381 ppm 0 381 ppm 50 mL 2.96% Cu2+ ppm in Initial Sample 2574 ppm 2574 ppm 2574 ppm 2574 ppm 0 2574 ppm 7.4 mL

1 2 3 Average RSD CI

The Beers Law limitations are grouped into three categories, namely: real deviations, chemical deviations and instrumental deviations. Real deviations are the fundamental limits of the Beers Law, since it the law only applies to dilute solutions. Solutions whose molarity is exceeding 0.01M are inapplicable with the Beers Law because the average distances of the molecules or ions are diminished to the extent that they affect each others charge distribution and absorption. This limit also applies to dilute solutions whose absorbers are in contact with a high concentration of other species of other species. Chemical deviations on the other hand are the errors that we are unaware of. These are the problems regarding the equilibrium of the solution. They are usually uncorrectable (Christian, 503-505). The last limitation is the instrumental deviation, which is just the problems with the spectrophotometer itself or the containing cell. Problems arise when the use of a monochromatic light is not observed, as differences in wavelength arise, the linearity deviates. The position of the cuvette also affects the retrieved data, the path length of light should be constant at all times. The last possible factor is the stray light, a radiation that is outside the nominal wavelength band chosen for determination. It usually comes from reflection of surfaces (Christian, 504-505) Better results could have been obtained if the experiment itself is replicated and proper handling and washing of cuvette and the use of spectrophotometer was implemented. The covering of the reagents and samples should also be observed to preserve the purity of the solution. Although there are restrictions to spectrophotometry, it is still widely used and has its own applications to various fields of science and technology. Environmental applications are mostly employed on analysis of water and wastewater. It is also used to determine the organic constituents of water and significant airborne pollutants. In the clinical field, spectrophotometry is applied in the analysis of drugs and narcotics. It is also used to determine the blood alcohol using the Breathalyzer test. All in all, the application of this experiment is mainly for the analysis of industrial processed samples (Harvey, 395-398). CONCLUSION The analysis of Cu2+ concentration using UV-Vis spectrophotometry resulted to very accurate and precise results. The analytical wavelength was found

Table 1 shows the calculated data from the analysis of the sample copper solution. The concentrations and absorbances of the three samples of working analyte are determined to be equal and consistent to 381ppm and 0.386 respectively. There would be no relative standard deviation and the confidence interval is just the concentration itself. The initial analytes concentration is determined to be consistent to 2574ppm, after multiplying working sample ppm to the dilution factor that is 50mL/7.4mL from the 7.4 aliquot being diluted to 50mL. There is also no RSD and the CI is just the concentration of itself. The error percentage from the theoretical value of 2500ppm is 2.96%, this error is cleary seen on the y-intercept value of the calibration curve. The y-intercept determines if there is any deviation on the linear relationship. In an ideal equation the y-intercept should be equal to zero, but the Beers Law has its own set of limitations and the working environment is not a vacuum that is strictly controlled. To completely understand the deviation, the limitations of Beers law and the errors in the experiment were presented and discussed in the next paragraphs. In this experiment, the analysis has some errors, clearly signified by the y-intercept and the percent error. The miscalculations could have resulted from the inconsistency of the purity of solution, dust may have settled in, which increases the absorbance of the analyte. Dirt and fingerprints could also be be present, this limits the transmittance. Stray light and iproper cleaning of cuvette also affects the absorbance. The other factors are the Beers Law limitations.

to be 596nm by scanning the most concentrated solution. The equation of the calibration curve was derived as y = 0.001x 0.005 (7). The concentration of the sample is 2574 ppm, this gives a very small deviation of 2.94%. The relative standard deviation is zero since the absorbances are all the same. The confidence interval is just the concentration of the analyte itself, which is 2574 ppm. The experiment could have yielded better results if replication of procedures and data was followed. There could be a great improvement if the working environment was maintained at standard conditions. However, the experiment still produced very accurate and precise results, which proves the efficiency of spectrophotometry as a mode of analysis and a best example of separation science. REFERENCES Brown, T. et al. 2004. Chemistry: The Central Science. Pearson. USA Harvey, David. 2000. Modern Analytical Chemistry. Mcgraw Hill. USA Skoog D.A., West D.M., Holler F.J., 1996. Fundamentals of analytical chemistry, 7th Ed. Saunders College Publishing, USA Jeffrey, G.H. 1989. Vogels Textbook Of Quantitative Chemical Analysis 5th Ed. Longman & Scientific Technical. USA Christian, G. 2004. Analytical Chemistry. John Wiley and Sons. USA