Determination of Iron in Beer

1. Prepare a table indicating how you prepared the standard solutions for the analysis of iron in beer. Tabulate the absorbances measured at 510 nm. Include in your table the relevant information for the beer sample (Note: question is repeated and bolded to set it apart from the answer below) Table 1. Data for the Beer’s Law Plot for Determination of Iron in Beer. (Final volume = 1000 µL) (note all column headings and numbers are centered) µL 1.00 x10-3 M 1 Fe2+ µL H2O µL 0.0100 M 2 PHE µL 20 % 3 ASC µL 20 % 4 NaOAc (in cuvet) 1 0 100 20 100 780 0.00 0.025 2 20 80 “ “ “ 2.00 0.225 3 40 60 “ “ “ 4.00 0.495 4 60 40 “ “ “ 6.00 0.665 5 80 20 “ “ “ 8.00 0.945 6 100 0 “ “ “ 10.00 1.075 7 -- 25 “ “ “ -- 0.095 9 -- 25 “ “ “ -- 0.092 9 -- 50 “ “ “ -- 0.167 10 -- 50 “ “ “ -- 0.163 Standard Curve 5 Fe2+1 10-5 M Absorbance 510 nm Beer (Note: some explanatory information is provided in footnotes) 1 2 3 4 5 Fe2(NH4)2(SO4)2 . 6 H2O was used as the primary standard for a 0.1987 M stock solution in 0.010 M HCl. A portion of this stock solution was diluted with 0.010 M HCl to achieve a 1.00 x 10-3 M solution used to prepare the standard solutions. PHE is 1,10-phenanthroline dissolved in 0.010 M HCl water. ASC is ascorbic acid, w/v %. NaOAc is sodium acetate, w/v %. Coors Lite beer, undiluted 1 2. Using your data for the standard iron solutions, construct two Beer’s law plots. For Figure 1 plot absorbance versus mg/mL iron(II); for Figure 2, plot absorbance versus M of iron(II). Attach these plots to the end of your report and any spreadsheet(s) used to construct the plots. Show calculations for the extinction coefficient below for Figure 2. Extinction coefficient: The extinction coefficient is found using Beer’s Law equation, A = ε c l, where ε is the extinction coefficient, c the concentration, and l the path length of the cuvet (1.00 cm): ε = slope of plot of A vs concentration in mol/L from standard plot = 0.108 x 105 M-1 cm-1 = 1.08 x 104 M-1 cm-1 See regression analysis for complete determination of extinction coefficient. 2 3. Determine the unknown iron in your beer sample by two methods. For Method I, use Figure 1 and read the iron concencentration in mg/mL in the cuvet from the plot directly. Convert it to mg/mL iron in your sample. For Method II, use the extinction coefficient determined from the slope of Figure 2 to find the unknown iron concentration in M in the cuvet. Convert it to the molarity of iron in your sample. Method I: Sample Calculation for Sample #7: (Note 2 sig. figs. are used because method is less accurate) blank correction ABS at 510 nm = 0.095 - 0.025 = 0.070 Iron concentration in cuvet = 0.33 x 10-3 mg/mL (Read directly from Figure 1) Iron concentration in sample = 0.33 x 10-3 mg/mL x 1000 µL --------25 µL = 13 x 10-3 mg/mL (volume correction Method II: for dilution into cuvet) The concentration of iron in an unknown sample is determined using Beer’s Law equation, A = ε c l, where the individual absorbance values are divided by the known extinction coefficient multipled by the path length Sample Calculation for Sample #7: ABS at 510 nm = 0.095 blank correction Iron in cuvet Iron in Beer = 0.095 - 0.025 ---------------------------------- = 0.0648 x 10-4 M =6.48 x 10-6 M 1.08 x 104 M-1 cm-1 (1.00 cm) (Carry extra digit - this is not final “answer”) -6 = 6.48 x 10 M x 1000 µL -----------25 µL = 2.6 x 10-4 M (Correct sig. figs. for final “answer”) (volume correction for dilution into cuvet) (Note: where multiple calculations are tabulated in a spreadsheet, it is not necessary to show all calculations. If all calculations are shown, they must be summarized in a table with proper headings. The summary table is placed BEFORE the calculations) 3 4. Compare the iron concentration in your beer sample determined by Method I and Method II. Note that you will have to convert mg/mL to M or visa versa. To be consistent, convert mg/mL to M. (carry extra digit here) Method I: 13.4 x 10-3 mg 1g ------------- x --------mL 103 mg Method II: 2.56 x 10-4 M x 103 mL ---------- x 1L 1 mole -------- = 2.4 x 10-4 M 55.85 g 2.5 ± 0.10 x 10-4 M Average Iron in sample: (sig figs for comparison limited by least accurate determination) Sample Method I: Method II: Average: Deviation ---------- = Average Iron value 2.4 x 10-4 M 2.6 x 10-4 M -------------2.5 x 10-4 M Deviation from average 0.10 x 10-4 M 0.10 x 10-4 M --------------0.10 x 10-4 M 0.10 x 10-4 M --------------- x100 % 2.5 x 10-4 M = 4.0 % The two methods give the same iron in the beer sample to within 4.0% 5. What are the two forms of iron that may be found in beer? Why is it likely to find one form and not the other? Iron has two oxidation states, Fe3+ and Fe2+. The Fe2+ ion is not likely to be present because it is readily oxidized by dissolved oxygen. Therefore the Fe3+ ion is likely to be the one present in beer. It is important to note that beer contains dissolved acids (lactic, citric, etc.) from the manufacturing process using yeast; these acids will chelate to the Fe3+ thereby preventing its precipitation from the beer as Fe2O3 (rust!). Since it taints the flavor of beer, Fe3+ levels are keep at a low level in beer. 4 6. Write the chemical reaction that is occurring between iron and 1,10-phenanthroline in this system? What is the function of the ascorbic acid and the sodium acetate? Iron (II) and 1,10-phenanthroline form a complex where three phenanthroline molecules surround the iron(II) ion forming a colored complex: Fe2+(aq) + 3 PHE º Fe(PHE)3(aq) The ascorbic acid acts as a reducing agent to reduce any Fe3+ that could be present to Fe2+ so that it can bind to the phenanthroline. The sodium acetate acts to maintain the pH around 4 to 5 which is optimal for the formation of the colored complex. (Note: the question has 3 parts that are clearly answered!) 7. How do you know that the absorbing species in the beer sample is iron(II)-phenanthroline complex and not some other metal ion-phenanthroline complex? The species giving the color with the beer sample was shown to be the iron(II)-phenanthroline complex after comparison of the visible spectra of the reagent mixtures of the beer sample and one of the standards (see Figure 3). Both spectra both had the same wavelength maximum and the same overall shape. They were not identical because the iron concentration in the beer sample was not the same as in the standard sample. 8. Discuss three sources of errors in your experiment. The major source of error is in pipetting technique. The pipetors are not calibrated and so that the volumes delivered may not be what is dialed in the window. Secondly, poor pipeting technique (pipeting from source at a distance, delivery from the pipet tip that is not at eye level, careless uptake and delivery in other ways, etc.). Finally, another source of error is inadequate mixing; solutions pipetted together do not necessarily mix thoroughly. (Note: provide a thoughtful analysis of errors in your experiment. Do not copy this paragraph into any of your reports!) 5 Standard Plot for the Determination of Iron Note: all calculations are performed in the spreadsheet Note: other reagents are left off of this spreadsheet Stock Solution of Iron Standard Final volume in cuvet MW Fe µL Fe2+ -3 Sample 1 2 3 4 5 6 Std 10 mg/mL in cuvet 0 20 40 60 80 100 0.00 1.12 2.23 3.35 4.47 5.58 0.0558 1.00 1000 55.85 -5 ABS 510 nm CORR ABS 510 nm 0.0 2.0 4.0 6.0 8.0 10.0 0.025 0.225 0.495 0.665 0.945 1.075 0.000 0.200 0.470 0.640 0.920 1.050 Fe2+ 10 M in cuvet CORR ABS 510 nm (Can also use trendline information) SUMMARY OUTPUT ABS vs M Regression Statistics Mult R 0.9966 2 R 0.9933 Adj R2 0.9916 Standard Error 0.0372 Obs 6 Intercept X Var 1 Coef 0.0052 0.1083 Std Err 0.0269 0.0044 X Variable 1 = Extinction Coefficient x 105 M-1 cm-1 0.108 (cannot ignore magnitude of x axis) or x 104 M-1 cm-1 1.08 (Beer's Law: A = ε c l) ε is the slope of A vs c with l = 1.00 cm Beer Undiluted1 µL ABS Sample 510 nm mg/mL 10-3 M µL g/mole Calc Iron Iron in Beer Iron in Beer 10-4 M in cuvet2 10-3 mg/mL 10-3 mg/mL Using Figure 1: Absorbance versus Fe 3+ mg/mL 7 8 9 10 1 2 25 25 50 50 0.095 0.092 0.167 0.163 0.070 0.067 0.142 0.138 0.33 0.32 0.71 0.68 AVE = 13 2.4 13 2.3 14 2.5 14 2.5 13 2.4 -3 - The beer required no predilution 10 mg/mL 10-4 M (cannot ignore magnitude) 2+ - Obtained from plot of Absorbance versus mg/mL Fe by reading directly from graph; fewer sig figs Using Figure 2: Absorbance versus Fe 2+ M Calc Iron in cuvet3 10-4 M 0.0648 0.0620 0.1315 0.1278 AVE = Iron in Beer 10-4 M 2.59 2.48 2.63 2.56 2.56 10-4 M (cannot ignore magnitude) 3 - Obtained from unknown absorbance and extinction coefficient, ε, and A = εcl 6 1.200 no box around plot 12 font caption (no title) correct significant figures 1.000 ABSORBANCE 0.800 0.600 Note: Draw best straight line through points going through (0,0). Find unknown ABS on y-axis, read + corresponding Fe3 in mg/mL on xaxis. Don't forget the order of magnitude. 0.400 0.200 Note: Correct superscripts and correct order of magnitude indicated 0.000 0.00 1.00 2.00 3.00 4.00 5.00 6.00 Fe2+ (10-3 mg/mL) Figure 1. Standard plot for the determination of iron using 1,10phenanthroline and ascorbic acid in sodium acetate solution. The absorbance is measured at 510 nm. Use "Print Preview" mode to check that caption wraps correctly 7 1.200 no box around plot 12 font caption (no title) correct significant figures 1.000 ABSORBANCE 0.800 0.600 0.400 Note: Draw best straight line through points going through (0,0). Find ε = ∆ x/ ∆ y using points as far apart as possible on the line . Don't forget the order of magnitude on the x-azis 0.200 0.000 0.00 Note: Correct superscripts and correct order of magnitude indicated 2.00 4.00 6.00 8.00 10.00 12.00 [Fe2+] (10-5 M) Figure 2. Standard plot for the determination of iron using 1,10-phenanthroline and ascorbic acid in sodium acetate solution. The absorbance is 2. measured at 510 nm. The extinction coefficient from these data is -4 -1 -1 1.08 x 10 M cm . Use "Print Preview" mode to check that caption wraps correctly 8 1.000 A 0.900 0.800 ABSORBANCE 0.700 0.600 0.500 B 0.400 0.300 0.200 0.100 0.000 450 475 500 525 550 575 600 625 WAVELENGTH (nm) Figure 3. (A) Spectrum of iron(II)-phenanthroline complex prepared from a standard iron sample. (B) Spectrum obtained from the analysis of a Lite Coors beer sample. 9 650