Efavirenz Capsules
DEFINITION
Efavirenz Capsules contain NLT 92.0% and NMT 108.0% of the labeled amount of efavirenz (C14H9ClF3NO2).
IDENTIFICATION
•  A. Infrared Absorption 197K
Sample solution:  Dissolve the contents of 1 Capsule in about 5 mL of acetonitrile by mixing on a vortex mixer. Allow to settle, remove about 3 mL of the solution, and centrifuge for about 5 min. Transfer 1–2 mL of supernatant to a clean suitable container, and evaporate to dryness under nitrogen. Mix 0.5–1 mg of the powder with 200 mg of potassium bromide.
•  B. Ultraviolet Absorption 197U
Solvent:  Acetonitrile
Standard solution:  10 µg/mL in Solvent
Sample solution:  Dissolve the contents of 1 Capsule in about 40 mL of Solvent by shaking for about 30 min. Pass through a suitable nylon or PVDF membrane filter, discarding the first 2 mL of filtrate, and dilute a portion with acetonitrile to a concentration of 10 µg/mL of efavirenz.
Acceptance criteria:  The UV absorption spectrum of the Sample solution exhibits maxima and minima at the same wavelengths as does the Standard solution.
ASSAY
•  Procedure
Diluent:  Acetonitrile and water (1:1)
Solution A:  Methanol, trifluoroacetic acid, and water (1:0.005:9). [Note—Use only freshly-opened trifluoroacetic acid, 6 months. ]
Solution B:  Methanol, trifluoroacetic acid, and water (9:0.005:1). [Note—Use only freshly-opened trifluoroacetic acid, 6 months. ]
Mobile phase:  See the gradient table below.
Time
(min)
Solution A
(%)
Solution B
(%)
0 60 40
16 50 50
23 35 65
28 30 70
29 20 80
31 20 80
32 60 40
40 60 40
Standard solution 1:  0.2 mg/mL of USP Efavirenz Related Compound B RS in Diluent
Standard solution 2:  5 mg/mL of USP Efavirenz RS in acetonitrile. [Note—Sonicate to dissolve before diluting to final volume. ]
Standard solution:  250 µg/mL of USP Efavirenz RS and 1 µg/mL of USP Efavirenz Related Compound B RS in Diluent prepared from Standard solution 2 and Standard solution 1, respectively. [Note—Store protected from light. For the HPLC analysis, it is recommended to use polypropylene vials, because degradation has been noted with certain brands made of glass. ]
Sample stock solution:  Transfer the contents of NLT 10 Capsules to a suitable container, and extract the contents in acetonitrile by mixing for about 30 min to obtain a quantitative solution equivalent to about 5 mg/mL of efavirenz. [Note—Store protected from light. ]
Sample solution:  Filter a portion of the Sample stock solution, and dilute the filtrate with Diluent to obtain a solution of about 250 µg/mL of efavirenz. [Note—Store protected from light. For the HPLC analysis, it is recommended to use polypropylene vials, because degradation has been noted with certain brands made of glass. ]
Chromatographic system 
Mode:  LC
Detector:  UV 250 nm
Column:  4.6-mm × 15-cm; packing L10
Column temperature:  40
Flow rate:  1.5 mL/min
Injection size:  35 µL
System suitability 
Sample:  Standard solution
Suitability requirements 
Resolution:  NLT 1.2 between efavirenz related compound B and efavirenz
Relative standard deviation:  NMT 2.0% for efavirenz
Analysis 
Samples:  Standard solution and Sample solution
Calculate the percentage of efavirenz (C14H9ClF3NO2) in the portion of Capsules taken:
Result = (rU/rS) × (CS/CU) × 100
rU== peak response of efavirenz from the Sample solution
rS== peak response of efavirenz from the Standard solution
CS== concentration of USP Efavirenz RS in the Standard solution (mg/mL)
CU== nominal concentration of efavirenz in the Sample solution (mg/mL)
Acceptance criteria:  92.0%–108.0%
PERFORMANCE TESTS
•  Dissolution 711
Medium:  1.0% (w/v) sodium lauryl sulfate in water; 900 mL. [Note—Do not deaerate. ]
Apparatus 2:  50 rpm, with helix sinker
Time:  45 min
Standard solution:  (L/900) mg/mL of USP Efavirenz RS in Medium, where L is the Capsule label claim in mg. A small volume of methanol, NMT 10% of the final volume, could be used to solubilize efavirenz. Dilute this solution with Medium to obtain a final concentration of about 0.01 mg/mL for Capsules labeled to contain 50 mg, or about 0.02 mg/mL for Capsules labeled to contain 100 mg or 200 mg.
Sample solution:  Pass a portion of the solution under test through a suitable filter of 0.45-µm pore size, and dilute with Medium to obtain a theoretical concentration similar to the Standard solution, assuming complete dissolution of the Capsule label claim.
Analytical wavelength:  UV 247 nm
Cell:  1 cm
Blank:  Medium
Analysis 
Samples:  Standard solution and Sample solution
Calculate the percentage of efavirenz dissolved:
Result = (AU/AS) × (CS/L) × D × V × 100
AU== absorbance of the Sample solution
AS== absorbance of the Standard solution
CS== concentration of the Standard solution (mg/mL)
L== label claim (mg/Capsule)
D== dilution factor of the Sample solution
V== volume of Medium, 900 mL
Tolerances:  NLT 80% (Q) of the labeled amount of efavirenz is dissolved.
•  Uniformity of Dosage Units 905: Meet the requirements
Procedure for content uniformity 
Standard solution:  10 µg/mL of USP Efavirenz RS in acetonitrile
Sample solution:  Transfer the contents of 1 Capsule into a suitable container, and dissolve in 40.0 mL of acetonitrile. Shake for about 30 min and pass through a suitable nylon or PVDF membrane filter. Dilute a portion of the filtrate to an efavirenz concentration of about 10 µg/mL.
Spectrometric conditions 
Mode:  UV absorption spectroscopy
Analytical wavelength:  UV 246 nm
Cell:  1 cm
Blank:  Acetonitrile
Analysis 
Samples:  Standard solution and Sample solution
Calculate the percentage of efavirenz (C14H9ClF3NO2) in the portion of Capsules taken:
Result = (AU/AS) × (CS/L) × V × D × 100
AU== absorbance of efavirenz from the Sample solution
AS== absorbance of efavirenz from the Standard solution
CS== concentration of USP Efavirenz RS in the Standard solution (mg/mL)
L== label claim (mg/Capsule)
V== volume of the Sample solution
D== dilution factor of the Sample solution
IMPURITIES
Organic Impurities 
•  Procedure
Diluent, Solution A, Solution B, Sample solution, and Chromatographic system:  Prepare as directed in the Assay.
System suitability solution:  Use the Standard solution prepared as directed in the Assay.
Standard solution:  1.25 µg/mL of USP Efavirenz RS and 0.005 µg/mL of USP Efavirenz Related Compound B RS in Diluent from the System suitability solution
System suitability 
Samples:  System suitability solution and Standard solution
Suitability requirements 
Resolution:  NLT 1.2 between efavirenz related compound B and efavirenz, System suitability solution
Relative standard deviation:  NMT 5.0% for efavirenz, Standard solution
Analysis 
Samples:  Standard solution and Sample solution
Calculate the percentage of any individual impurity in the portion of Capsules taken:
Result = (rU/rS) × (CS/CU) × (1/F) × 100
rU== peak response of any individual impurity (degradation product) from the Sample solution
rS== peak response of efavirenz from the Standard solution
CS== concentration of USP Efavirenz RS in the Standard solution (mg/mL)
CU== nominal concentration of efavirenz in the Sample solution (mg/mL)
F== relative response factor (see Impurity Table 1)
Acceptance criteria 
Individual impurities:  See Impurity Table 1. [Note—Disregard any peak less than 0.05%. ]
Total impurities:  NMT 0.50%. [Note—Include only the degradation products in the calculation of the total impurities. ]
Impurity Table 1
Name Relative
Retention
Time
Relative
Response
Factor
Acceptance
Criteria,
NMT (%)
Efavirenz aminoalcohol (degradation product)a 0.48 0.26 0.25
Efavirenz ethene analogb 0.93 *
Efavirenz pent-3-ene-1-yne (cis)c 1.16 *
Efavirenz pent-3-ene-1-yne (trans)d 1.16 *
Efavirenz penteneynee 1.16 *
Efavirenz pentyne analogf 1.2 *
Methylefavirenzg 1.28 *
Efavirenz aminoalcohol methyl carbamateh 1.33 *
N-Benzylefavirenzi 1.8 *
Efavirenz benzoylaminoalcoholj 1.9 *
Quinoline analog (degradation product)k 1.45 2.0 0.20
Efavirenz aminoalcohol ethyl carbamatel 1.53 *
Unidentified impurity 1.60 *
Efavirenz aminoalcohol bis(ethoxycarbonyl)m 1.63 *
Unidentified impurity 2.1 *
Cyclobutenylindole analogn 2.18 *
Any other individual degradation product 1.0 0.20
a   (S)-2-(2-Amino-5-chlorophenyl)-4-cyclopropyl-1,1,1-trifluorobut-3-yn-2-ol.
b   (S,E)-6-Chloro-4-(2-cyclopropylvinyl)-4-(trifluoromethyl)-2H-3,1-benzoxazin-2-one.
c   (S,E)-6-Chloro-4-(pent-3-en-1-ynyl)-4-(trifluoromethyl)-2H-3,1-benzoxazin-2-one.
d   (S,Z)-6-Chloro-4-(pent-3-en-1-ynyl)-4-(trifluoromethyl)-2H-3,1-benzoxazin-2-one.
e   (S)-6-Chloro-4-(3-methylbut-3-en-1-ynyl)-4-(trifluoromethyl)-2H-3,1-benzoxazin-2-one.
f   (S)-6-Chloro-4-(pent-1-ynyl)-4-(trifluoromethyl)-2H-3,1-benzoxazin-2-one.
g   (S)-6-Chloro-4-{[(2RS,2RS)-2-methylcyclopropyl]ethynyl}-4-(trifluoromethyl)-2H-3,1-benzoxazin-2-one.
h   (S)-Methyl 4-chloro-2-(4-cyclopropyl-1,1,1-trifluoro-2-hydroxybut-3-yn-2-yl)phenylcarbamate.
i   (S)-6-Chloro-4-(cyclopropylethynyl)-1-(4-methoxybenzyl)-4-(trifluoromethyl)-2H-3,1-benzoxazin-2-one.
j   (S)-N-[4-Chloro-2-(4-cyclopropyl-1,1,1-trifluoro-2-hydroxybut-3-yn-2-yl)phenyl]-4-methoxybenzamide.
k   6-Chloro-2-cyclopropyl-4-(trifluoromethyl)quinoline.
l   (S)-Ethyl 4-chloro-2-(4-cyclopropyl-1,1,1-trifluoro-2-hydroxybut-3-yn-2-yl)phenylcarbamate.
m   (S)-Ethyl 4-chloro-2-[4-cyclopropyl-2-(ethoxycarbonyloxy)-1,1,1-trifluorobut-3-yn-2-yl]phenylcarbamate.
n   Ethyl 5-chloro-2-cyclobutenyl-3-(trifluoromethyl)-1H-indole-1-carboxylate.
*   For information purposes only. These are process impurities monitored in the drug substance and are not included in the total impurities.
ADDITIONAL REQUIREMENTS
•  Packaging and Storage: Store in well-closed containers at controlled room temperature.
•  USP Reference Standards 11
USP Efavirenz RS Click to View Structure
USP Efavirenz Related Compound B RS
(S,E)-6-Chloro-4-(2-cyclopropylvinyl)-4-(trifluoromethyl)-2H-3,1-benzoxazin-2-one.
    C14H11ClF3NO2        317.69
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