Spectrophotometry in the Visible Region á851ñ —

Solution: 10µg per mL.

Medium: pH7.5phosphate buffer,prepared by mixing 21volumes of 0.15Mdibasic sodium phosphate and 4volumes of 0.15Mmonobasic potassium phosphate.

Ratio: A405/A630,between 3.0and 3.9

Copper stock solution and Standard solutions—Prepare as directed in the test for Content of total copper.

Test solution— Transfer about 100mg of Chlorophyllin Copper Complex Sodium,accurately weighed,to a 150-mLconical flask,add 75mLof water,and shake by mechanical means for about 3minutes.Adjust with 1Nhydrochloric acid to a pHof 3.0,transfer the suspension thus obtained to a 100-mLvolumetric flask,dilute with water to volume,and mix.Filter this suspension,discarding the first 10mLof the filtrate.Use the clear filtrate as the Test solution.

Procedure— Proceed as directed for Procedurein the test for Content of total copper.Calculate the percentage of ionic copper in the portion of Chlorophyllin Copper Complex Sodium taken by the formula: in which the terms are as defined therein:not more than 0.25%is found,calculated on the dried basis.

Copper stock solution— Transfer 1.000g of copper to a 1000-mLvolumetric flask,dissolve in 20mLof nitric acid,dilute with 0.2Nnitric acid to volume,and mix.This solution contains 1000µg of copper per mL.Store in a polyethylene bottle.

Standard solutions— Pipet 5.0mLof Copper stock solutioninto a 500-mLvolumetric flask,dilute with water to volume,and mix.Transfer 5.0,10.0,15.0,and 20.0mL,respectively,of this solution to separate 100-mLvolumetric flasks,dilute the contents of each flask with water to volume,and mix.These Standard solutionscontain 0.5,1.0,2.0,and 3.0µg of copper per mL,respectively.

Test solution— Transfer about 100mg of previously dried Chlorophyllin Copper Complex Sodium,accurately weighed,to a Kjeldahl flask.Add 2.0mLof sulfuric acid,1.0mLof nitric acid,and 1.0mLof hydrogen peroxide,and carefully heat under a fume hood until a light green color is obtained.[NOTE—If the solution has any hint of a brown tint,continue to add 0.5-mLportions of nitric acid until a green color is obtained.]Cool,transfer the contents quantitatively to a 1000-mLvolumetric flask with several portions of water,dilute the contents of the flask with water to volume,and mix.Transfer 10.0mLof this solution to a 50-mLvolumetric flask,dilute with water to volume,and mix.

Procedure— Concomitantly determine the absorbances of the Standard solutionsand the Test solutionat the copper emission line of 324.8nm with an atomic absorption spectrophotometer (see Spectrophotometry and Light-Scattering á851ñ)equipped with a copper hollow-cathode lamp and an air–acetylene flame,using water as the blank.Plot the absorbances of the Standard solutionsversus the concentration,in µg per mL,of copper,and draw the straight line best fitting the four plotted points.From the graph so obtained,determine the concentration,C,in µg per mL,of copper in the Test solution.Calculate the percentage of copper in the portion of Chlorophyllin Copper Complex Sodium taken by the formula: in which Wis the weight,in mg,on the dried basis,of Chlorophyllin Copper Complex Sodium taken to prepare the Test solutionnot less than 4.25%is found.

Standard stock solution— Dissolve 254.2mg of sodium chloride,previously dried at 105for 2hours,in 50mLof water,transfer to a 1000-mLvolumetric flask,dilute with water to volume,and mix.Each mLof this solution contains 100µg of sodium.

Standard solutions— Transfer to each of five 100-mLvolumetric flasks 10mLof a solution of a nonionic wetting agent (1in 500).Dilute the contents of one of the flasks with water to volume to provide a blank.To the remaining flasks add,respectively,2.5,5.0,10.0,and 15.0mLof the Standard stock solution,dilute with water to volume,and mix.These Standard solutionscontain 2.5,5.0,10.0,and 15.0µg of sodium per mL,respectively.

Test solution— Transfer about 100mg of Chlorophyllin Copper Complex Sodium,accurately weighed,to a 1000-mLvolumetric flask,add 100mLof a solution of nonionic wetting agent (1in 500)and 400mLof water,and shake by mechanical means for about 5minutes.Dilute with water to volume,and mix.

Standard graph— Set the flame photometer for maximum transmission at a wavelength of about 589nm.Adjust the instrument to zero transmittance with the blank.Adjust the instrument to 100%transmittance with the most concentrated of the Standard solutions.Read the percentage of transmittance of the other Standard solutions,and plot the percentage of transmittance versus the concentration,in µg per mL,of sodium.

Procedure— Adjust the instrument as directed under Standard graph,read the percentage of transmittance of the Test solution,and from the Standard graphread the concentration,C,in µg per mL,of sodium in the Test solution.Calculate the percentage of sodium in the portion of Chlorophyllin Copper Complex Sodium taken by the formula: in which Wis the weight,in mg,of Chlorophyllin Copper Complex Sodium taken to prepare the Test solution:between 5%and 7%,calculated on the dried basis,is found.