4826 Polyethylene / Official Monographs USP 36

Add 120 mL of acetonitrile, mix, and degas. [NOTE—Protect Mobile phase—Dilute 40.0 mL of Salt solution with water
the Mobile phase from air to prevent absorption of carbon to 1000 mL. Make adjustments if necessary (see System Suit-
dioxide.] Make adjustments if necessary (see System Suitabil- ability under Chromatography 〈621〉).
ity under Chromatography 〈621〉). Increasing the proportion Standard preparation—Transfer about 360 mg of USP Pol-
of buffer solution decreases the retention times of the yethylene Glycol 3350 RS, accurately weighed, to a 500-mL
analytes. volumetric flask, add 20.0 mL of Salt solution and about
Internal standard solution—Dissolve a suitable quantity of 250 mL of water, dissolve by swirling, dilute with water to
ammonium bromide in water to obtain a solution having a volume, and mix. This Standard preparation contains about
concentration of about 2.2 mg per mL. 0.72 mg of polyethylene glycol 3350 per mL.
Standard preparation—To a 100-mL volumetric flask trans- Assay preparation—Use the Assay preparation, prepared as
fer about 246 mg of sodium chloride (4.2 mEq), previously directed in the Assay for potassium and sodium. This solution
dried at 105° for 2 hours and accurately weighed, and contains about 0.72 mg of polyethylene glycol 3350 per mL.
about 682 mg of anhydrous sodium sulfate (9.6 mEq), pre- Chromatographic system (see Chromatography 〈621〉)—
viously dried at 105° for 2 hours and accurately weighed, [NOTE—Use peak heights where peak responses are indi-
dilute with water to volume, and mix. Transfer 5.0 mL of cated.] The liquid chromatograph is equipped with a refrac-
this solution to a 500-mL volumetric flask, add 10.0 mL of tive index detector maintained at 34 ± 0.5°, a 7.8-mm ×
Internal standard solution, dilute with water to volume, and 4.5-cm guard column containing packing L25, and a 7.8-
mix. Filter this solution through a 0.5-µm or finer porosity mm × 30-cm analytical column containing packing L25 and
filter, and store the filtrate in a suitable glass container. This maintained at ambient temperature. The flow rate is about
Standard preparation contains about 24.6 µg of sodium chlo- 1 mL per minute. Chromatograph the Standard preparation
ride (0.00042 mEq of chloride) and about 68.2 µg of so- as directed for Procedure: the relative standard deviation for
dium sulfate (0.00096 mEq of sulfate) per mL. replicate injections is not more than 1.5%. [NOTE—Maintain
Assay preparation—Use the Assay preparation prepared as column backpressure at less than 1000 pounds per square
directed in the Assay for potassium and sodium. This solution inch. Backpressure may be reduced by cleaning the frits in
contains about 0.042 mEq of chloride and 0.096 mEq of the guard column or by replacing the guard column. Base-
sulfate per mL. line drift may be reduced by maintaining strict control of
Chromatographic system (see Chromatography 〈621〉)—The ambient temperature, by insulating the lines, the Mobile
liquid chromatograph is equipped with a conductivity de- phase reservoir, and the columns, and by increasing the
tector, a 4-mm × 5-cm guard column containing packing time of equilibration.]
L23, and a 4-mm × 30-cm analytical column maintained at Procedure—Separately inject equal volumes (about 20 µL)
35 ± 1° containing packing L23. The flow rate is about of the Standard preparation and the Assay preparation into
0.9 mL per minute. Chromatograph the Standard prepara- the chromatograph, record the chromatograms, and meas-
tion as directed for Procedure: the relative retention times are ure the responses for the major peaks. Calculate the con-
about 0.25 for chloride, 0.4 for bromide, and 1.0 for sulfate, tent, in g, of polyethylene glycol 3350 per L of constituted
the resolution, R, between the chloride and bromide peaks Oral Solution taken by the formula:
is not less than 1.5 and between the bromide and sulfate
peaks is not less than 4.5. [NOTE—Maintain column back- 500(C / 6)(rU / rS)
pressure at less than 1000 pounds per square inch. Back-
pressure may be reduced by changing the in-line filters and in which C is the concentration, in mg per mL, of polyethyl-
frits in the columns. Column efficiency may be improved by ene glycol 3350 in the Standard preparation, and rU and rS
backflushing the analytical column with 50 mL of the buffer are the polyethylene glycol 3350 peak responses obtained
solution used to prepare the Mobile phase.] from the Assay preparation and the Standard preparation, re-
Procedure—[NOTE—Use peak heights where peak re- spectively.
sponses are indicated.] Separately inject equal volumes
(about 10 µL) of the Standard preparation and the Assay
preparation into the chromatograph, record the chromato-
grams, and measure the responses for the major peaks. Cal-
Polymyxin B Sulfate
.

culate the mEq of chloride per L of constituted Oral Solution

taken by the formula: Polymyxin B, sulfate.
(500 / 58.44)(C / 6)(RU / RS) Polymyxin B sulfate [1405-20-5].

in which 58.44 is the molecular weight of sodium chloride,

» Polymyxin B Sulfate is the sulfate salt of a kind
C is the concentration, in µg per mL, of sodium chloride in of polymyxin, a substance produced by the
the Standard preparation, and RU and RS are the peak re- growth of Bacillus polymyxa (Prazmowski) Migula
sponse ratios of chloride to bromide obtained from the As- (Fam. Bacillaceae), or a mixture of two or more
say preparation and the Standard preparation, respectively. such salts. It has a potency of not less than 6000
Calculate the mEq of sulfate per L of constituted Oral Solu-
tion taken by the formula: Polymyxin B Units per mg, calculated on the
dried basis.
(500 / 71.02)(C / 6)(RU / RS)
Packaging and storage—Preserve in tight, light-resistant
in which 71.02 is one-half of the molecular weight of so- containers.
dium sulfate, C is the concentration, in µg per mL, of anhy- Labeling—Where packaged for prescription compounding,
drous sodium sulfate in the Standard preparation, and RU the label states the number of Polymyxin B Units in the
and RS are the peak response ratios of sulfate to bromide container and per milligram, that it is not intended for man-
obtained from the Assay preparation and the Standard prepa- ufacturing use, that it is not sterile, and that its potency
ration, respectively. cannot be assured for longer than 60 days after opening.
Assay for polyethylene glycol 3350— Where it is intended for use in preparing injectable or other
sterile dosage forms, the label states that it is sterile or must
Salt solution—Prepare a solution in water containing be subjected to further processing during the preparation of
0.35 mg of sodium chloride, 0.18 mg of potassium chloride, injectable or other sterile dosage forms.
0.40 mg of sodium bicarbonate, 1.37 mg of anhydrous so-
dium sulfate, and 0.88 mg of ammonium bromide per mL. USP Reference standards 〈11〉—
USP Polymyxin B Sulfate RS
USP 36 Official Monographs / Polymyxin 4827

Identification— 90.0 percent and not more than 120.0 percent of

A: Liquid Chromatographic Identification Test— the labeled amount of polymyxin B.
Mobile phase—Prepare a mixture of 0.1 M tribasic so-
dium phosphate and acetonitrile (77:23), and adjust with Packaging and storage—Preserve in Containers for Sterile
phosphoric acid to a pH of 3.0. Make adjustments if neces- Solids as described under Injections 〈1〉, protected from light.
sary (see System Suitability under Chromatography 〈621〉. Labeling—Label it to indicate that where it is administered
Standard solution—Prepare a solution of USP Polymyxin B intramuscularly and/or intrathecally, it is to be given only to
Sulfate RS in Mobile phase having a concentration of about patients hospitalized so as to provide constant supervision
3.5 mg per mL. Protect this solution from light. by a physician.
Test solution—Prepare a solution of Polymyxin B Sulfate in USP Reference standards 〈11〉—
Mobile phase having a concentration of about 3.5 mg per USP Polymyxin B Sulfate RS
mL. Protect this solution from light. Constituted solution—At the time of use, it meets the
Chromatographic system (see Chromatography 〈621〉)— requirements for Constituted Solutions under Injections 〈1〉.
The liquid chromatograph is equipped with a 212-nm de- Thin-layer chromatographic identification test
tector and a 4.6-mm × 25-cm column that contains 5-µm 〈201BNP〉: meets the requirements.
packing L1. The flow rate is about 1 mL per minute. Pyrogen—It meets the requirements of the Pyrogen Test
Procedure—Separately inject equal volumes (about 10 µL) 〈151〉, the test dose being 1.0 mL per kg of a solution in
of the Standard solution and the Test solution into the chro- pyrogen-free saline TS containing 20,000 Polymyxin B Units
matograph, and record the chromatograms. The chromato- per mL.
gram obtained from the Test solution corresponds qualita- Sterility 〈71〉—It meets the requirements when tested as
tively to that obtained from the Standard solution, exhibiting directed for Membrane Filtration under Test for Sterility of the
a major peak corresponding to polymyxin B1 and peaks at Product to be Examined.
relative retention times of about 0.5 (polymyxin B2) and 0.6
(polymyxin B3). Particulate matter 〈788〉: meets the requirements for
small-volume injections.
B: Dissolve 2 mg in 5 mL of water, add 5 mL of 2.5 N
sodium hydroxide, mix, and add 5 drops of cupric sulfate Residue on ignition 〈281〉: not more than 5.0%, the
solution (1 in 100), shaking after the addition of each drop: charred residue being moistened with 2 mL of nitric acid
a reddish violet color is produced. and 5 drops of sulfuric acid.
C: A solution (1 in 20) meets the requirements of the Heavy metals, Method II 〈231〉: not more than 0.01%.
tests for Sulfate 〈191〉. Other requirements—It meets the requirements for pH
pH 〈791〉: between 5.0 and 7.5, in a solution containing and Loss on drying under Polymyxin B Sulfate. It also meets
5 mg per mL. the requirements for Uniformity of Dosage Units 〈905〉 and
for Labeling under Injections 〈1〉.
Loss on drying 〈731〉—Dry about 100 mg, accurately
weighed, in a capillary-stoppered bottle in vacuum at 60° Assay—
for 3 hours: it loses not more than 7.0% of its weight. Assay preparation 1 (where it is represented as being in a
Content of phenylalanine—Transfer about 0.375 g of single-dose container)—Constitute Polymyxin B for Injection
Polymyxin B Sulfate, accurately weighed, to a 100-mL volu- in a volume of water, accurately measured, corresponding
metric flask, dissolve in and dilute with 0.1 N hydrochloric to the volume of solvent specified in the labeling. Withdraw
acid to volume, and mix. Measure the absorbances of this all of the withdrawable contents, using a suitable hypoder-
solution at the maxima at about 264 nm (A264), 258 nm mic needle and syringe, and dilute quantitatively with Buffer
(A258), and 252 nm (A252), and the absorbances at 280 nm No. 6 to obtain a solution containing a convenient number
(A280) and 300 nm (A300). Calculate the percentage of phen- of Polymyxin B Units per mL.
ylalanine in the portion of Polymyxin B Sulfate taken by the Assay preparation 2 (where the label states the quantity
formula: of polymyxin B in a given volume of constituted solution)—
Constitute 1 container of Polymyxin B for Injection in a vol-
(9.4787/W)(A258 – 0.5A252 + 0.5A264 – 1.84A280 + 0.8A300) ume of water, accurately measured, corresponding to the
volume of solvent specified in the labeling. Dilute an accu-
in which W is the weight, in g, of Polymyxin B Sulfate rately measured volume of the constituted solution quantita-
taken: it contains between 9% and 12% of phenylalanine, tively with Buffer No. 6 to obtain a solution containing a
calculated on the dried basis. convenient number of Polymyxin B Units per mL.
Other requirements—If for prescription compounding, it Procedure—Proceed as directed under Antibiotics—Micro-
meets the requirements for Residue on ignition under bial Assays 〈81〉, using an accurately measured volume of
Polymyxin B for Injection. Where the label states that Assay preparation diluted quantitatively with Buffer No. 6 to
Polymyxin B Sulfate is sterile, it meets the requirements for yield a Test Dilution having a concentration assumed to be
Sterility Tests 〈71〉 and, where intended for injectable dosage equal to the median dose level of the Standard.
forms, for Pyrogen under Polymyxin B for Injection. Where the
label states that Polymyxin B Sulfate must be subjected to
further processing during the preparation of injectable dos-
age forms, it meets the requirements for Pyrogen under
Polymyxin B for Injection. Polymyxin B Sulfate and Bacitracin
.

Assay—Proceed with Polymyxin B Sulfate as directed under Zinc Topical Aerosol

Antibiotics—Microbial Assays 〈81〉.
» Polymyxin B Sulfate and Bacitracin Zinc Topical
Aerosol contains the equivalent of not less than
90.0 percent and not more than 130.0 percent of
Polymyxin B for Injection the labeled amounts of polymyxin B and
.

bacitracin.
» Polymyxin B for Injection contains an amount Packaging and storage—Preserve in pressurized contain-
of Polymyxin B Sulfate equivalent to not less than ers, and avoid exposure to excessive heat.