DETERMINATION OF COPPER (II) CONCENTRATION BY COLORIMETRIC METHOD

H.M. TOLENTINO INSTITUTE OF BIOLOGY, COLLEGE OF SCIENCE UNIVERSITY OF THE PHILIPPINES, DILIMAN, QUEZON CITY, PHILIPPINES
DATE SUBMITTED: SEPTEMBER 27, 2011

ABSTRACT
The experiment intended to introduce spectrometric methods of analysis, specifically Beers law. It also affirmed the proper use of a UV-Vis spectrophotometer. A calibration curve was constructed in order to determine the concentration of the unknown sample by using 0.0, 2.0, 4.0, 6.0, 8.0 and 10.0 mL portions of the working standard solution diluted to 50.0 mL and 10.0 mL of concentrated ammonia was added to each. The solutions to be analyzed were put in a 4.0-5.0 mL cuvette with the reagent blank being the first one put in the UV-Vis spectrophotometer. The analytical wavelength ( wavelength of maximum absorbance) was determined first by putting the 500 ppm solution and the machine displayed the wavelength. The unknown solution was determined by substituting it to the equation of the best fit curve and the concentration was 239 ppm Cu2+. It was then multiplied by the aliquot factor and resulted to 1195 ppm which is the concentration of the unknown stock sample. The standard deviation of the unknown sample was also determined and yielded as 2.391. The small deviation implied that the constructed calibration curve was precise (having R2 as 0.9999) and with a very small value of the y-intercept (0.0012) indicating small real deviations. The results revealed the effectiveness of spectrometric analyses in determining concentrations of ions.

INTRODUCTION
The subject of spectroscopy is the interaction of radiation with matter. Spectroscopic analytical methods are based on measuring the amount of radiation produced or absorbed by molecular or atomic species of interest.[1] The modern era of spectroscopy began when Sir Isaac Newton observed the prism spectrum of the sun in 1672. In 1802, William Hyde Wollaston noted dark lines while observing the solar spectrum with a spectroscope. These lines were later described in detail by Joseph Fraunhofer, beginning in 1817 who gave the prominent lines letters starting with A at the red end of the spectrum. The Fraunhofer line were explained later by Gustav Robert Kirchoff and Robert Willhelm Bunsen in 1859 and 1860 and concluded that the Fraunhofer A and B lines were due to the potassium in the suns atmosphere while D lines were due to sodium. Kirchoff and Bunsen are thus credited with discovering the use of spectroscopy for chemical analysis.[1] The idea of spectroscopy is that all atoms or molecules absorb energy from at least one region of the spectrum of electromagnetic radiation. Spectroscopic methods are classified according to the region of the electromagnetic spectrum involved in the measurement. When absorption is from the UV/ visible region of the spectrum (200-700), transitions

occur between electronic energy levels, and these electronic transitions form the physical basis for the techniques used in spectrophotometry. [2] Absorption of energy in molecules only happen when the incident photon has an energy precisely equal to the difference in energy between two allowed states, the photon promoting the transition of an electron from the lower to the higher energy state. Before another photon can be absorbed, the excited state must lose this energy and revert to the ground state, commonly at a speed of 10-12 seconds and occurs by loss of energy to vibrations and rotations within the same molecule and by collision, to other molecules.[2] In other words, the process of absorption transfers energy to the molecule and results in a decrease in the intensity of the incident electromagnetic radiation. Energy is lost to the environment as heat. Absorption of the radiation thus attenuates the beam in accordance with the absorption law or the Beer-Bouger-Lambert law.[1] Bouger in 1729 and Lambert in 1760 recognized that when electromagnetic energy is absorbed, the power of the transmitted energy decreases exponentially. Later, in 1852, Beer and Bernard each stated that a similar law holds for the dependence of transmittance on the concentration. The laws were combined and is now what we call the Beers law.[3]

The use of spectroscopy in chemical analyses was proven to be effective. Some of the elements were discovered using spectroscopic methods. Typical applications of these methods are usually found in the field of biochemistry. Complex organic molecules are analyzed through the use of these methods. The use colors of the visible spectrum also enable scientists to determine the composition of the atmosphere of planets and stars without leaving earth. The experiment aimed to determine the concentration of Cu2+ ions in a given unknown solution using spectrophotometric methods of analysis. The standard solution used was Cu(NO3)2 3H2O. A UV-Vis spectrophotometer was the instrument used to determine the absorbance of the solutions. The original procedure required the determination of the analytical wavelength (the wavelength at which the solution absorbs strongly). But due to improved capabilities of the modern instrument, the manual determination of the analytical wavelength was no longer needed since the machine automatically finds it. Using samples from the standard solution of known Cu2+ concentrations, a calibration curve was made, having a plot of absorbance against concentration(in ppm). The equation of the curve was determined. This will be the working equation in determining the Cu2+ concentration of the unknown solution. The purpose of this investigation is to be familiar with spectroscopic methods of analyses specifically the use and application of Beer s law in calculations. It also aims one to be familiar on how a spectrophotometer operates and the concept behind it.

RESULTS AND DISCUSSION

The absorption of light by a sample is described by Beer s law. Consider the absorption of radiation as in figure 1.

Incident radiation of radiant power P0 passes through a cell containing a solution of an absorbing species at concentrations c and pathlength b, and the transmitted radiation has radiant power P. The transmittance T of the solution is the fraction of incident radiation transmitted by the solution and decays exponentially with pathlength and concentration as shown in equation (1). The absorbance A of a solution is related to the transmittance in a logarithmic manner as shown in equation (2). This implies that an increase in the absorbance will result in a decrease in transmittance. The use of absorbance in calculations is also more convenient thus, modern spectrometers have linear absorbance scales or a computer that calculates absorbance from measured quantities.[1] Combining equations (1) and (2) will result in equation (3) which is the common form of Beers law. Note the presence of the constant a in equations (1) and (3), it is called the absorptivity and is dependent on the wavelength and the nature of the absorbing material. In an absorption spectrum, the absorbance varies with wavelength in direct proportion to a.[3] If c is expressed in g/L, the units of a is g-1c-1L since absorbance is a unitless quantity. If c is in mol/L, instead of a, molar absorptivity is used and has units g-1mol-1L. Thus, A= bc The UV-Vis spectrophotometer works by measuring the transmitted incident radiation after passing on a solution placed in a cuvette. The cuvette used is a rectangular glass, open at the top with transparent faces on both ends of the pathlength (entry and exit sides of radiation) while translucent at the other two parallel faces. It has a capacity of approximately 4.0-5.0 mL. The machine has a built-in function that determines the analytical wavelength of the solution. From that, the plot of absorbance against wavelength was no longer needed. In order to determine the Cu2+ concentration of the unknown solution, a calibration curve was constructed first using different amounts (0.00, 2.00, 4.00, 6.00, 8.00 and 10.00 mL) from a working standard of concentration 2500 ppm Cu2+. 10 mL of concentrated NH3 was put in each solution to ensure that the Cu2+ ions are fully dissociated throughout the solution and that they all absorb the radiation. The curve was made by plotting the (A) absorbance measured at 623 nm( analytical wavelength) against concentration of the standard Cu2+ solution (ppm Cu2+) for the given series of standard solutions.

(1) (2) (3)

0.5 Absorbance (A) 0.4 0.3 0.2 0.1 0 0 200 400 600

SUMMARY AND CONCLUSIONS

Since there was only one trial for the unknown sample, the standard deviation cannot be applied, instead, the standard deviation of the unknown sample from the calibration curve was taken and resulted to a relatively small value. This showed that the analysis was valid and the resulting concentration of 239 ppm for the sample while 1195 ppm for the stock solution, was acceptable. Even though there were deviations in the calibration curve, the calculated value showed only a slight deviation. Possible sources of error can be traced down in the solution preparation. Proper amounts of the working standard might not have been transferred quantitatively. Instrumental error cannot be determined since the apparatus was assumed to be in its best condition.

Concentration (c, ppm Cu) Figure 1. Calibration curve for obtaining concentration by absorbance measurements, generated through spreadsheet. The resulting equation of the best fit line is y = 0.000894x + 0.0012 with R2 of 0.999889913. Beers law states that absorbance is directly proportional to concentration where b is a constant or directly proportional to pathlength at a fixed concentration. On the actual experiment, b was held constant (1 cm). The concentration of the unknown solution was calculated by using the equation of the standard curve. The equation of the standard curve has the form y = mx + b, which is the linear form of the Beers law (A = abc). Theoretically, the b in the equation of the curve must be equal to zero. But due to the limitations of the Beers law, a non-zero value of b exists. These small deviations are called real deviations. Other limitations of the law are instrumental deviations, result of the method to make absorbance measurements and chemical deviations which are results of chemical changes within the solution.[4] The Beers law is also considered as a limiting law since it only describes the absorption behavior of dilute solutions.[4] That is why the equation of the standard curve was used in determining the concentration of the unknown solution instead of the normal form of the Beers law. Only one trial of analysis was performed for the unknown solution. The calculated concentration of the unknown analyzed solution was 239 ppm Cu2+. The value is valid since it fell under the domain of the calibration curve (100-500 ppm). Multiplying it by the aliquot factor 5, it will become 1195 ppm Cu2+, the concentration of the unknown stock solution. Since only one trial was performed, the standard deviation of the sample concentration to the calibration curve was determined. The standard deviation of the unknown concentration is 2.391, the concentration is reported as 239 2.391 ppm Cu2+. The other team had a concentration of 273 ppm (1365 ppm of the stock) from an absorbance of 0.249. The mean was taken to be 256 ppm or 1280 ppm of the stock solution.

REFERENCES
[1] Skoog D., D. West, et.al. 2000. Analytical Chemistry, An Introduction7th edition. Brooks/Cole [2] Harris, D. A., C. L. Bashford, eds. 1987. Spectrophotometry and Spectroflourimetry: A Practical approach.IRL Press Limited [3] Christian G., D. et. Al. 2004. Analytical Chemistry. 6th edition. John Wiley & Sons,Inc. [4] Skoog D., West D., et al. 2010. Fundamentals of Analytical Chemistry, 8th Edition. Brooks/Cole

APPENDIX Raw Data

Concentration of standard Cu2+ 100 ppm 200 ppm 300 ppm 400 ppm 500 ppm Absorbance 0.091 0.181 0.266 0.361 0.448

Working Equations  e.g.

 

Where: Sc = standard deviation of the unknown concentration Sr = standard deviation about regression; standard error of estimate m = slope of the calibration curve M = number of replicate analyses of unknown N = number of measurements yc = sample yave = average of y values Sxx = sum of deviations from the mean xave value Syy = sum of deviations from the mean yave value