Experiment 25 The Fertilizer Project Analysis of Phosphorus in Fertilizer Spectrophotometric Construct a Calibration Curve 1 Obtain 20 mL of a standard stock solution of 5 00x10E 3 0 into a dry 50 mL beaker 2 Create a working solution by using a Mohr pipet to obtain 3 00mL of the standard stock solution into a 250 mL volumetric flask and adding enough water to dilute the solution to the 250 mL mark 3 Use a volumetric pipet to place 5 00 mL into a 10 mL volumetric flask add 4mL of heteropoly blue solution and dilute to the 10mL mark 4 Pour all of the solution into a medium test tube and label correctly 5 Repeat steps 3 and 4 with 1 0 2 0 3 0 and4 0 working stock solutions 6 Develop a reagent blank by adding 4mL of the heteropoly blue solution to the 10mL volumetric flask add distilled water to the mark pour into a medium test tube and label it a A reagent blank must be made as a reference solution to ensure no PO4 3 exists in the distilled water in addition to make sure no other constituents of the solution contribute to the absorbance 7 Develop the blue color by placing the test tubes in boiling water for 10 minutes 8 Allow the test tubes to cool to room temperature 9 Tilt each test tube to mix the solution with the water that has condensed in the test tube 10 Use a spectrometer to calculate the absorbance spectra for a block the light path and the reference the reagent blank solution 11 Obtain an absorbance spectra for each colored solution ensuring the arrow is pointing toward the light source the cuvet does not contain air bubbles the cuvet is filled and using the same cuvet for each of your colored solutions and rinsing the cuvet with a small amount of the next solution to be measured before filling it up to be measured 12 Print or record your results 13 Using the software Absorbance Series to plot your results from step 12 in one graph 14 If any point deviate far from the line of best fit you must remeasure that point 15 Determine your wavelength and print your results 16 Select Beer s Law Plot for the Analysis menu and enter absorbance values and concentrations for each colored solution a This plot is your calibration curve for A vs PO43 Analyzing the Sample 1 Determine the amount of fertilizer needed to make 250 mL of a solution 2 Use the KH2PO4 fertilizer solid to prepare a sample solution a Place the calculated amount in our case 36g of the fertilizer into a 250 mL volumetric flask 3 4 5 6 7 b Add 3 00mL of this solution into another 250mL volumetric flask creating your working solution Prepare three colored solutions using three different volumes of the working solution 4mL of the heteropoly blue solution and water into 3 medium sized test tubes Create another reagent blank solution by placing 3mL of the diluted fertilizer solution and water into a 10mL volumetric flask and filling the rest with water Record and save the absorption spectrum for each solution and generate a Beer s Law plot for the sample data Use the equation from the calibration curve to determine the amount of Phosphate in the each colored sample of the diluted fertilizer by assuming there is one PO43in each heteropoly blue formula unit see calculation 3 Convert the amount of phosphate found in the solution to P2O5 and compare it to the expected value see calculation 7 8 1 Amount of fertilizer sample needed 100gfertilizer 250L 5 10 3 30 97 35gfertilizer 11gm 2 Calculation of P in colored sample for calibration curve P KH2PO4 5 00x10 3 mol 3mL 1 or 3 or 6 mLaliquot x x 1L 250mL 10mLsol 3 Mass of P in 250 mL fertilizer stock solution xxxx10 6 molP 10mL colored calibrationcurve x P incoloredsolution 1L xxxmL aliquot 4 250 mL working solution mole P in colored solution x P in working solution 3 mL mole P in working solution x 250L mol Pin fertilizer sample solution 5 6 30 97 g P mole P in fertilizer solution x g P in fertilizer sample solution 1 mol P 7 Calculation of P2O5 in fertilizer 141 9 g P2O5 xxx g P x xxx g P2O5 61 94 g P 8 g P2O5 P2O5 x x 100 g fertilizer Gravimetric Method Procedure 1 Weigh out 0 1 0 01 g of KH2PO4 on an analytical balance 2 Dissolve the sample in about 10 mL of deionized water in a 50 mL beaker 3 Add 10 mL of the 0 4 M MgSO4 solution 4 Using a disposable plastic pipet slowly add 1 mL 6M NH3 dropwise while stirring after adding all of the NH3 touch the stirring rod to a piece of red litmus paper 5 Repeat this adding 1 mL of NH3 at a time until the litmus turns blue when the solution contacts it 6 Then add 0 5 mL more of NH3 This will precipitate insoluble Mg NH4 PO4 6H2O Let the solution stand about 15 min to complete the precipitation 7 Filter off the precipitate using suction filtration Wash the precipitate with two 5mL portions if iso propyl alcohol 8 Suck air through the filter for about twenty minutes to help dry the precipitate and then carefully scrape it onto a pre weighed watch glass Be sure that you get all of the precipitate 9 Let the precipitate dry in air for half an hour and weigh the watch glass with precipitate If your precipitate is not dry the weight will continue to change with time 10 Repeat steps 1 9 for a second trial 11 Measure mass of product You will use this to determine the of P2O5 see calculation 3 12 Repeat steps 1 11 replacing KH2PO4 in step 1 with 0 91g of fertilizer see calculation 1 1 Amount of fertilizer needed for the analysis 100 g of fertilizer x 0 1 g P 0 91 g of 11 g of P Fertilzer 2 Mass of Phosphorous in original sample 1 mol Mg NH 4 PO4 6H2O 3 1 mol P Measured g Mg NH 4 PO4 6H2O 245 5 g Mg NH 4 PO4 6H2O 1 mol Mg NH 4 PO4 6H2O 2 5 3 of P O in Fertilizer g P2O5 141 94 mol g P 2 O5 g P g 61 94 P mol g P2O5 100 P2O5 g of Fertilzer measured