Anticoagulant Citrate Phosphate Dextrose Adenine Solution, chemical structure, molecular formula, Reference Standards

Anticoagulant Citrate Phosphate Dextrose Adenine Solution

»Anticoagulant Citrate Phosphate Dextrose Adenine Solution is a sterile solution of Citric Acid,Sodium Citrate,Monobasic Sodium Phosphate,Dextrose,and Adenine in Water for Injection.It contains,in each 1000mL,not less than 2.11g and not more than 2.33g of monobasic sodium phosphate (NaH2PO4·H2O);not less than 30.30g and not more than 33.50g of dextrose (C6H12O6·H2O);not less than 19.16g and not more than 21.18g of total citrate,expressed as citric acid,anhydrous (C6H8O7);not less than 6.21g and not more than 6.86g of sodium (Na);and not less than 0.247g and not more than 0.303g of adenine (C5H5N5).It contains no antimicrobial agents.

Prepare Anticoagulant Citrate Phosphate Dextrose Adenine Solution as follows:

Citric Acid (anhydrous)	2.99g
Sodium Citrate (dihydrate)	26.3g
Monobasic Sodium Phosphate (monohydrate;NaH2PO4·H2O)	2.22g
Dextrose (monohydrate)	31.9g
Adenine (C5H5N5)	0.275g
Water for Injection,a sufficient quantity,to make	1000mL

Dissolve the ingredients,and mix.Filter the solution until clear,place immediately in suitable containers,and sterilize.

If desired,3.27g of monohydrated citric acid may be used instead of the indicated amount of anhydrous citric acid;23.06g of anhydrous sodium citrate may be used instead of the indicated amount of dihydrated sodium citrate;1.93g of anhydrous monobasic sodium phosphate may be used instead of the indicated amount of monohydrated monobasic sodium phosphate;and 29.0g of anhydrous dextrose may be used instead of the indicated amount of monohydrated dextrose.

Packaging and storage— Preserve in single-dose containers,of colorless,transparent,Type Ior Type IIglass,or of a suitable plastic material (see Transfusion and Infustion Assemblies and Similar Medical Devices á161ñ).

Labeling— Label it to indicate the number of mLof solution required per 100mLof whole blood or the number of mLof solution required per volume of whole blood to be collected.

USP Reference standards á11ñ— USP Adenine RS.USP Endotoxin RS.

Bacterial endotoxins á85ñ— It contains not more than 5.56USP Endotoxin Units per mL.

pHá791ñ: between 5.0and 6.0.

Chloride á221ñ— A10-mLportion shows no more chloride than corresponds to 0.50mLof 0.020Nhydrochloric acid (0.0035%).

Other requirements— It meets the requirements under Injections á1ñ.

Assay for total citrate—

Standard preparations— Dissolve a suitable quantity of citric acid,previously dried at 90

for 3hours and accurately weighed,in water to obtain a solution having a known concentration of about 1.0mg of anhydrous citric acid per mL.Further pipet quantities of 8,9,10,11,and 12mLof this stock standard into separate 100-mLvolumetric flasks,dilute with water to volume,and mix.

Assay preparation— Pipet 5mLof Solution into a 1000-mLvolumetric flask,dilute with water to volume,and mix.

Procedure— Pipet 1mLof the Assay preparation,each Standard preparation,and water into separate test tubes.To each tube add 1.3mLof pyridine,and mix by swirling.To one tube at a time add 5.7mLof acetic anhydride,and mix,using a rotary vortex stirrer.Immediately place in a water bath maintained at 31±1.0

,and allow the color to develop for 33±1minutes.Determine the absorbance against the reference blank in 1-cm cells at 425nm,taking care to measure the absorbance of each solution at the same elapsed time from mixing.Calculate the total citrate content,in mg per mL,of the Solution taken by the formula:

0.2C,

in which Cis the concentration,in µg per mL,of anhydrous citric acid read from the standard curve.

Assay for total phosphate [expressed as monobasic sodium phosphate monohydrate (NaH2PO4·H2O)]—

Acid solution— Dilute 75mLof sulfuric acid to 200mLwith water.

Ammonium molybdate solution— Dissolve 25g of ammonium molybdate in 300mLof water,and add 200mLof Acid solution.

Standard preparation— Dissolve about 0.44g of monobasic potassium phosphate,previously dried at 105

for 4hours and accurately weighed,in water to make 1000mL,and mix.Pipet 10mLof this solution into a 100-mLvolumetric flask,dilute with water to volume,and mix to obtain a solution having a known concentration of about 0.01mg of phosphorus (P)per mL.

Assay preparation— Pipet 10mLof Solution into a 500-mLvolumetric flask,dilute with water to volume,and mix.

Procedure— Pipet 5-,10-,and 15-mLportions of the Standard preparationand 10mLof the Assay preparationinto separate 50-mLvolumetric flasks.Treat the contents of each flask as follows.Add 2.5mLof Ammonium molybdate solution,and mix.Add rapidly,and in order,2.5mLof hydroquinone solution (1in 200)and 2.5mLof sodium sulfite solution (1in 10),both prepared fresh daily.Dilute with water to volume,mix,and allow to stand at room temperature for 30±5minutes.Determine the absorbances against water in 1-cm cells at 660nm with a suitable spectrophotometer.Plot the absorbances against the mg of phosphorus in the portions of Standard preparationtaken.Calculate the quantity,in mg,of NaH2PO4·H2Oin each mLof the Solution taken by the formula:

(137.99/30.9738)(5W),

in which 137.99is the molecular weight of monobasic sodium phosphate monohydrate;30.9738is the atomic weight of phosphorus;and Wis the weight,in mg,of Pin the 10mLof Assay preparationtaken,read from the Standard curve.

Assay for sodium—

Lithium diluent solution— Transfer 1.04g of lithium nitrate to a 1000-mLvolumetric flask,add a suitable nonionic surfactant,then add water to volume,and mix.This solution contains 15mEq of lithium per L.

Standard preparation— Transfer 8.18g of sodium chloride,previously dried at 105

for 2hours and accurately weighed,to a 1000-mLvolumetric flask,dilute with water to volume,and mix.This solution contains 140mEq of sodium per L.Transfer 50µLof this solution to a 10-mLvolumetric flask,dilute with Lithium diluent solutionto volume,and mix.

Assay preparation— Pipet 25mLof Solution into a 50-mLvolumetric flask,dilute with water to volume,and mix.Transfer 50µLof this solution to a 10-mLvolumetric flask,dilute with Lithium diluent solutionto volume,and mix.

Procedure— Using a suitable flame photometer,adjusted to read zero with Lithium diluent solution,concomitantly determine the sodium flame emission readings for the Standard preparationand the Assay preparationat the wavelength of maximum emission at about 589nm.Calculate the quantity,in g,of Na in 1000mLof Anticoagulant Citrate Phosphate Dextrose Adenine Solution taken by the formula:

2(8.18)(22.99/58.44)(RU/RS),

in which 8.18is the weight,in g,of sodium chloride taken to make the Standard preparation;22.99is the atomic weight of sodium;58.44is the molecular weight of sodium chloride;and RUand RSare the sodium emission readings obtained from the Assay preparationand the Standard preparation,respectively.

Assay for dextrose— Tare a clean,medium-porosity filtering crucible containing several carborundum boiling chips or glass beads.Pipet 50mLof freshly mixed alkaline cupric tartrate TSinto a 400-mLbeaker.Add the boiling chips or glass beads from the tared crucible,45mLof water,and 5.0mLof Solution to the beaker.Heat the beaker and contents over a burner that has been adjusted to cause boiling of the solution to start in 3.5to 4minutes.Boil the solution for 2minutes,accurately timed,and filter immediately through the tared crucible,taking care to transfer all of the boiling chips or glass beads to the crucible.Wash the precipitate with hot water and 10mLof alcohol.Dry the crucible and contents at 110

to constant weight.Perform a blank determination,and make any necessary correction.Each mg of cuprous oxide precipitate obtained is equivalent to 0.496mg of C6H12O6·H2O.

Assay for adenine—

Mobile phase— Dissolve 3.45g of ammonium dihydrogen phosphate in 950mLof water in a 1000-mLvolumetric flask,add 10mLof glacial acetic acid,dilute with water to volume,mix,pass through a membrane filter having a 1-µm or finer porosity,and degas.

Standard preparations— Dissolve accurately weighed quantities of USP Adenine RSin dilute hydrochloric acid (1in 120)in three separate volumetric flasks,dilute with the dilute hydrochloric acid solution to volume,and mix to obtain Standard preparations having known concentrations of about 0.25,0.275,and 0.30mg of adenine per mL,respectively.Protect from light.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 254-nm detector and a 4-mm ×30-cm stainless steel column that contains packing L9.The flow rate is about 2.0mLper minute.Prepare a solution containing USP Adenine RSand purine,each at about 0.275mg per mL,in dilute hydrochloric acid (1in 120),and chromatograph not less than four injections (about 20µL)of this solution:the relative standard deviation of the peak response of adenine is not more than 2.5%,the relative standard deviation of the retention time of adenine is not more than 2.0%,and the resolution of adenine and purine is not less than 3.0.

Procedure— Separately inject equal volumes (about 20µL)of the Solution and the Standard preparations,record the chromatograms,and measure the responses for the major peaks.Plot the responses against the concentrations,in mg of USP Adenine RSper mLof the Standard preparations.Calculate the quantity,in mg,of C5H5N5in each mLof the Solution taken as the value read directly from the Standard curve corresponding to the response obtained from the portion of the Anticoagulant Citrate Phosphate Dextrose Adenine Solution chromatographed.

Auxiliary Information— Staff Liaison:Radhakrishna S Tirumalai,Scientist

Expert Committee:(BBP)Blood and Blood Products

USP28–NF23Page 166

Phone Number:1-301-816-8339