Piperacillin and Tazobactam for Injection
DEFINITION
Piperacillin and Tazobactam for Injection contains amounts of Piperacillin Sodium and Tazobactam Sodium equivalent to NLT 90.0% and NMT 110.0% of the labeled amounts of piperacillin (C23H27N5O7S) and tazobactam (C10H12N4O5S), the labeled amounts representing proportions of piperacillin to tazobactam of 8:1. It may contain small amounts of a suitable buffer and stabilizer.
IDENTIFICATION
•  The retention times of the major peaks of the Sample solution correspond to those of the Standard solution, as obtained in the Assay.
ASSAY
•  Procedure
[Note—Refrigerate the Standard solution and the Sample solution immediately after preparation and during analysis, using a refrigerated autosampler set at 5 ± 3. The solutions should be analyzed within 24 h of preparation. ]
Solution A:  Phosphoric acid and water (1:4)
Solution B:  Dilute the contents of one vial of tetrabutylammonium hydrogen sulfate ion pairing reagent with water to 1 L.
Mobile phase:  Acetonitrile and Solution B (1:3), adjusted with Solution A to a pH of 3.8
Diluent:  Acetonitrile and water (1:3)
Standard stock solution A:  0.06 mg/mL of USP Tazobactam Related Compound A RS in Diluent
Standard stock solution B:  0.5 mg/mL of USP Tazobactam RS in Diluent
Standard stock solution C:  1.0 mg/mL of USP Piperacillin RS in acetonitrile and Diluent (1:24). [Note—Dissolve first in acetonitrile, using about 4% of the final volume, and dilute with Diluent to volume. ]
System suitability solution:  0.006 mg/mL of tazobactam related compound A from Standard stock solution A and 0.025 mg/mL of tazobactam from Standard stock solution B in Diluent
Standard solution:  0.025 mg/mL of tazobactam from Standard stock solution B and 0.2 mg/mL of piperacillin from Standard stock solution C in Mobile phase
Sample solution:  Equivalent to 0.025 mg/mL of tazobactam and 0.2 mg/mL of piperacillin from Piperacillin and Tazobactam for Injection in Mobile phase
Chromatographic system 
Mode:  LC
Detector:  UV 210 nm
Column:  4.6-mm × 15-cm; 3-µm packing L11
Autosampler temperature:  5 ± 3
Flow rate:  1 mL/min
Injection size:  20 µL
System suitability 
Samples:  System suitability solution and Standard solution
Suitability requirements 
Resolution:  NLT 3 between tazobactam related compound A and tazobactam, System suitability solution
Tailing factor:  NMT 1.8 for tazobactam and piperacillin, Standard solution
Relative standard deviation:  NMT 2% for tazobactam and piperacillin, Standard solution
Analysis 
Samples:  Standard solution and Sample solution
Calculate the percentages of C23H27N5O7S and C10H12N4O5S in the portion of Piperacillin and Tazobactam for Injection taken:
Result = (rU/rS) × (CS/CU) × P × 100
rU== peak response of piperacillin or tazobactam from the Sample solution
rS== peak response of piperacillin or tazobactam from the Standard solution
CS== concentration of USP Piperacillin RS or USP Tazobactam RS in the Standard solution (mg/mL)
CU== nominal concentration of piperacillin or tazobactam in the Sample solution (mg/mL)
P== potency of piperacillin or tazobactam in USP Piperacillin RS or USP Tazobactam RS, respectively (mg/mg)
Acceptance criteria:  90.0%–110.0%
PERFORMANCE TESTS
•  Uniformity of Dosage Units 905: Meets the requirements
IMPURITIES
Organic Impurities 
•  Procedure
[Note—Refrigerate the Standard solution and the Sample solution immediately after preparation and during analysis, using a refrigerated autosampler set at 5 ± 3. The solutions should be analyzed within 24 h of preparation. ]
Mobile phase, Diluent, System suitability solution, Standard solution, Sample solution, Chromatographic system, and System suitability:  Proceed as directed in the Assay.
Analysis 
Samples:  Standard solution and Sample solution
Calculate the percentage of each impurity in the portion of Piperacillin and Tazobactam for Injection taken:
Result = (rU/rS) × (CS/WU) × P × D × (1/F) × 100
rU== peak response of each impurity from the Sample solution
rS== response of piperacillin from the Standard solution
CS== concentration of USP Piperacillin RS in the Standard solution (mg/mL)
WU== weight of product used to prepare the Sample solution (mg)
P== potency of USP Piperacillin RS (mg/mg)
D== dilution factor of the Sample solution
F== relative response factor (see Impurity Table 1)
Acceptance criteria 
Individual impurities:  See Impurity Table 1.
Total impurities:  NMT 5.0%. [Note—Total impurities does not include piperacillin related compound A. ]
Impurity Table 1
Name Relative
Retention
Time
Relative
Response
Factora
Acceptance Criteria,
NMT (%)a
Tazobactam related compound Ab 0.12 0.75 1.0
Tazobactam 0.25
Piperacillin impurity 4c 0.31 1.0 1.0
Piperacillin impurity 1d,e 0.36 1.0 1.0
Piperacillin related compound Ae,f 0.51 0.56 5.0
Piperacillin related compound Cg 0.55 1.0 1.0
Piperacillin impurity 5c 0.62 1.0 1.0
Piperacillin impurity 6c 0.67 1.0 1.0
Piperacillin 1.0
Any individual unspecified impurity 1.0 1.0
a   Calculated relative to the peak area of piperacillin.
b   (2S,3S)-2-Amino-3-methyl-3-sulfino-4-(1H-1,2,3-triazol-1-yl)butyric acid.
c   Specified unidentified impurities.
d   (4S)-2-{[2-(4-Ethyl-2,3-dioxopiperazine-1-carboxamido)-2-phenylacetamido]methyl}-5,5-dimethylthiazolidine-4-carboxylic acid.
e   This compound has two epimers that usually co-elute but that may be separated as a result of minor changes in the chromatographic conditions.
f   (2R,4S)-2-{(1R)-Carboxy[2-(4-ethyl-2,3-dioxopiperazine-1-carboxamido)-2-phenylacetamido]methyl}-5,5-dimethylthiazolidine-4-carboxylic acid.
g   (2R,4S)-3-Acetyl-2-{(1R)-carboxy[2-(4-ethyl-2,3-dioxopiperazine-1-carboxamido)-2-phenylacetamido]methyl}-5,5-dimethylthiazolidine-4-carboxylic acid.
SPECIFIC TESTS
•  Bacterial Endotoxins Test 85: It contains NMT 0.08 USP Endotoxin Unit in a portion equivalent to 1 mg of a mixture of piperacillin and tazobactam (0.89 and 0.11 mg, respectively).
•  Sterility Tests 71: Meets the requirements
•  Particulate Matter in Injections 788: Meets the requirements
•  pH 791: 5.0–7.0, in a solution containing the equivalent of 40 mg/mL of piperacillin
•  Other Requirements: It meets the requirements under Injections 1.
ADDITIONAL REQUIREMENTS
•  Packaging and Storage: Preserve as described in Containers for Sterile Solids under Injections 1, Packaging. Store at controlled room temperature.
•  Labeling: Label it to indicate its sodium content.
•  USP Reference Standards 11
USP Endotoxin RS
USP Piperacillin RS Click to View Structure
USP Tazobactam RS Click to View Structure
USP Tazobactam Related Compound A RS Click to View Structure
(2S,3S)-2-Amino-3-methyl-3-sulfino-4-(1H-1,2,3-triazol-1-yl)butyric acid.
    C7H12N4O4S         248.26
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USP35–NF30 Page 4335
Pharmacopeial Forum: Volume No. 37(2)