Printed on: Tue Aug 03 2021, 07:34:44 AM Official Status: Currently Official on 03-Aug-2021 DocId: 1_GUID-F45751C8-CB8D-4930-8240-DDE6C466DC36_5_en-US

(EST)
Printed by: Le Tran Official Date: Official as of 01-May-2020 Document Type: USP @2021 USPC
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Suitability requirements
Xylazine Tailing factor: NMT 2.0
Relative standard deviation: NMT 2.0%
Analysis
Samples: Standard solution and Sample solution
Calculate the percentage of xylazine (C12H16N2S) in the
portion of Xylazine taken:

Result = (r U/r S) × (C S/C U) × 100

rU = peak response from the Sample solution

rS = peak response from the Standard solution
CS = concentration of USP Xylazine RS in the Standard
solution (mg/mL)
CU = concentration of Xylazine in the Sample solution
C12H16N2S 220.33
(mg/mL)
4H-1,3-Thiazin-2-amine, N-(2,6-dimethylphenyl)-5,6-
dihydro-; Acceptance criteria: 98.0%–102.0%
5,6-Dihydro-2-(2,6-xylidino)-4H-1,3-thiazine [7361-61-7].
IMPURITIES
DEFINITION • RESIDUE ON IGNITION á281ñ: NMT 0.1%
Xylazine contains NLT 98.0% and NMT 102.0% of xylazine • LIMIT OF 3-AMINO-1-PROPANOL
(C12H16N2S). Standard solution: 0.5 mg/mL of 3-amino-1-propanol in

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IDENTIFICATION methanol
Sample solution: 100 mg/mL of Xylazine in methanol.
Change to read: Sonicate to dissolve.
Chromatographic system
• A. ▲SPECTROSCOPIC IDENTIFICATION TESTS á197ñ, Infrared ci (See Chromatography á621ñ, General Procedures, Thin-Layer
Spectroscopy: 197A or 197K▲ (CN 1-May-2020) Chromatography.)
Mode: TLC
Change to read: Adsorbent: 0.25-mm layer of chromatographic silica gel
Application volume: 5 µL
• B. ▲SPECTROSCOPIC IDENTIFICATION TESTS á197ñ, Developing solvent system: Alcohol and ammonium
Ultraviolet-Visible Spectroscopy: 197U▲ (CN 1-May-2020)
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hydroxide (80:20)
Sample solution: 5 µg/mL in 0.1 N hydrochloric acid Spray reagent: 2 mg/mL of ninhydrin in alcohol
Acceptance criteria: Meets the requirements Analysis
ASSAY Samples: Standard solution and Sample solution
• PROCEDURE Develop the chromatogram in a saturated
Solution A: Dissolve 3.03 g of sodium 1-heptanesulfonate chromatographic chamber containing the Developing
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in 800 mL of water, adjust with 2 N sulfuric acid to a pH of solvent system until the solvent front has moved
3.0, and dilute with water to 1000 mL. Pass through a filter three-fourths of the length of the plate. Remove the plate
of 0.5-µm or finer pore size. from the chromatographic chamber, mark the solvent
Solution B: Acetonitrile front, and air-dry the plate. Spray the plate with the Spray
Diluent: Solution A and Solution B (50:50) reagent, and immediately heat the plate in an oven at
Mobile phase: See Table 1. Return to original conditions, 105°. When the spots are visible, remove the plate from
and equilibrate the system. the oven, and allow to cool.
Acceptance criteria: The intensity of the spot for 3-amino-
Table 1 1-propanol from the Sample solution is not greater than that
of the spot for 3-amino-1-propanol from the Standard
Time Solution A Solution B
(min) (%) (%) solution (0.5%).
• LIMIT OF ACETONE AND ISOPROPYL ALCOHOL
0 70 30 Proceed as directed in Residual Solvents á467ñ.
5 70 30 Acceptance criteria
Acetone: NMT 0.02%
10 60 40 Isopropyl alcohol: NMT 0.2%
15 60 40 • ORGANIC IMPURITIES
Solution A, Solution B, and Diluent: Prepare as directed
in the Assay.
Standard solution: 0.4 mg/mL of USP Xylazine RS in Diluent Mobile phase: See Table 2. Return to original conditions,
Sample solution: 0.4 mg/mL of Xylazine in Diluent and equilibrate the system.
Chromatographic system
(See Chromatography á621ñ, System Suitability.) Table 2
Mode: LC
Detector: UV 226 nm Time Solution A Solution B
(min) (%) (%)
Column: 3.9-mm × 30-cm; packing L1
Flow rate: 1 mL/min 0 75 25
Injection volume: 10 µL 8 75 25
System suitability
Sample: Standard solution 35 30 70

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Printed by: Le Tran Official Date: Official as of 01-May-2020 Document Type: USP @2021 USPC
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Table 2 (continued) CS = concentration of USP Xylazine RS in the Standard

Time Solution A Solution B solution (mg/mL)
(min) (%) (%) CU = concentration of Xylazine in the Sample solution
40 30 70
(mg/mL)
F = relative response factor (see Table 3)
Standard solution: 0.008 mg/mL of USP Xylazine RS in Acceptance criteria: See Table 3.
Diluent, prepared by diluting the Standard solution from the
Assay Table 3
Sample solution: Transfer 100 mg of Xylazine into a 10-mL Relative Relative Acceptance
volumetric flask, add 5.0 mL of Solution B, and swirl to Retention Response Criteria,
dissolve. Add 4 mL of Solution A, and swirl. Dilute with Name Time Factor NMT (%)
Solution A to volume, and mix. 2,6-Dimethylaniline 0.8 1.4 0.5
Chromatographic system
(See Chromatography á621ñ, System Suitability.) Xylazine 1.0 — —
Mode: LC Specified unknown 1.3 2.8 0.5
Detector: UV 205 nm
Column: 4.6-mm × 25-cm; packing L7 (use a guard 2,6-Dimethylphenyl iso-
column) thiocyanate 2.0 2.7 0.5
Flow rate: 1 mL/min Any individual
Injection volume: 10 µL unspecified —
System suitability impurity 1.0 0.5
Sample: Standard solution

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Total impurities — — 1.0
Suitability requirements
Tailing factor: NMT 1.5
SPECIFIC TESTS
Relative standard deviation: NMT 5.0%
• LOSS ON DRYING á731ñ
Analysis
Analysis: Dry under vacuum at 60° for 4 h.
Samples: Standard solution and Sample solution
Calculate the percentage of each impurity in the portion of
Xylazine taken:
ci Acceptance criteria: NMT 0.5%
ADDITIONAL REQUIREMENTS
• PACKAGING AND STORAGE: Preserve in tight containers.
Result = (r U/r S) × (C S/C U) × (1/F) × 100 Store at 25°, excursions permitted between 15° and 30°.
• LABELING: Where it is intended for veterinary use only, the
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rU = peak area of any individual impurity from the label so states.
Sample solution that is not present in the • USP REFERENCE STANDARDS á11ñ
chromatogram of the Diluent USP Xylazine RS
rS = peak area of xylazine from the Standard solution
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