Ritonavir

This article is compiled based on the United States Pharmacopeia (USP) – 2025 Edition

Issued and maintained by the United States Pharmacopeial Convention (USP)

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C₃₇H₄₈N₆O₅S₂ 720.94 

2,4,7,12-Tetraazatridecan-13-oic acid, 10-hydroxy-2-methyl-5-(1-methylethyl)-1-[2-(1-methylethyl)-4-thiazolyl]-3,6-dioxo-8,11-bis(phenylmethyl)-5- thiazolylmethyl ester [5S-(5R*,8R*,10R*,11R*)]-; 

5-Thiazolylmethyl [(αS)-α-[(1S,3S)-1-hydroxy-3-[(2S)-2-[3-[(2-isopropyl-4-thiazolyl)methyl]-3-methylureido]3-methylbutyramido]-4- phenylbutyl]phenethyl]carbamate CAS RN®: 155213-67-5; UNII: O3J8G9O825. 

1 DEFINITION 

Ritonavir contains NLT 97.0% and NMT 102.0% of ritonavir (C₃₇H₄₈N₆O₅S₂), calculated on the anhydrous basis. 

2 IDENTIFICATION 

Change to read: 

A. Spectroscopic Identification Tests 〈197〉, Infrared Spectroscopy: 197K

B. The retention time of the major peak of the Sample solution is within 2% of the retention time of the major peak of the Standard solution, as obtained in the Assay. 

3 ASSAY 

Procedure 

Solution A: 4.1 mg/mL of monobasic potassium phosphate in water 

Solution B: Acetonitrile, tetrahydrofuran (inhibitor-free), n-butanol, and Solution A (18:8:5:69) 

Mobile phase: Solution B 

Diluent: Acetonitrile and Solution A (1:1) 

Standard stock solution: 2.0 mg/mL of USP Ritonavir RS in Diluent. [Note—This solution may be kept for 5 days if refrigerated.] Standard solution 1: 0.10 mg/mL of USP Ritonavir RS from the Standard stock solution diluted with Diluent 

Standard solution 2: 0.025 mg/mL of USP Ritonavir RS from Standard solution 1 diluted with Diluent 

Sample solution: 0.025 mg/mL of Ritonavir in Diluent 

Chromatographic system 

(See Chromatography 〈621〉, System Suitability.) 

Mode: LC 

Detector: UV 240 nm 

Column: 4.6-mm × 15-cm; 3-µm packing L26 

Column temperature: 60° 

Flow rate: 1 mL/min 

Injection volume: 50 µL 

Run time: 40 min 

System suitability 

Sample: Standard solution 2 

Suitability requirements 

Capacity factor, k′: NLT 13 

Column efficiency: NLT 5000 theoretical plates 

Tailing factor: 0.8–1.2 

Relative standard deviation: NMT 2.0% 

Analysis 

Samples: Standard solution 2 and Sample solution 

Calculate the percentage of ritonavir (C₃₇H₄₈N₆O₅S₂) in the portion of Ritonavir taken: 

Result = (r_U/r_S) × (C_S/C_U) × 100 

r_U = peak response from the Sample solution 

r_S = peak response from the Standard solution 

C_S = concentration of USP ritonavir RS in the Standard solution (mg/mL)  

C_U = concentration of ritonavir in the Sample solution (mg/mL)

Acceptance criteria: 97.0%–102.0% on the anhydrous basis 

4 IMPURITIES 

Residue on Ignition 〈281〉: NMT 0.2%, determined on 1.0 g 

Organic Impurities 

Ritonavir is alkali sensitive. All glassware should be prerinsed with distilled water before use to remove residual detergent contamination. 

Solution A, Solution B, Mobile phase, Diluent, Standard stock solution, and Standard solution 1: Prepare as directed in the Assay. Solution C: Acetonitrile, tetrahydrofuran (inhibitor-free), n-butanol, and Solution A (47:8:5:40) 

Mobile phase: See Table 1. 

Table 1 

Identity solution: 1 mg/mL of USP Ritonavir Related Compounds Mixture RS in Diluent 

Standard solution 2: 5 µg/mL of USP Ritonavir RS from Standard solution 1 in Diluent. [Note—This is stable for 48 h.] Sample solution: 1 mg/mL of Ritonavir in Diluent 

Chromatographic system 

(See Chromatography 〈621〉, System Suitability.) 

Mode: LC 

Detector: UV 240 nm 

Column: 4.6-mm × 15-cm; 3-µm packing L26 

Column temperature: 60° 

Flow rate: 1 mL/min 

Injection volume: 50 µL 

Run time 

Standard solution 2: 40 min 

System suitability 

Samples: Identity solution and Standard solution 2 

[Note—See Table 2 for relative retention times.] 

Resolution: NLT 1.0 between hydroxyritonavir and hydantoin aminoalcohol peaks, Identity solution Peak-to-valley ratio: NLT 1 for ritonavir and the 4-hydroxy isomer, Identity solution 

Capacity factor, k′: NLT 13, Standard solution 2 

Column efficiency: NLT 5000 theoretical plates, Standard solution 2 

Tailing factor: 0.8–1.2, Standard solution 2 

Relative standard deviation: NMT 3.0%, Standard solution 2 

Analysis 

Samples: Diluent, Identity solution, Standard solution 2, and Sample solution 

Calculate the percentage of each impurity in the portion of Ritonavir taken: 

Result = (r_U/r_S) × (C_S/C_U) × (1/F) × 100 

r_U = peak response of each impurity from the Sample solution 2

r_S = peak response from the Standard solution 2

C_S = concentration of Standard solution 2 (mg/mL)  

C_U = concentration of ritonavir in the Sample solution (mg/mL)

F = relative response factor (see Table 2) 

Acceptance criteria: See Table 2. 

Table 2 

a Ureidovaline is [N-methyl[(2-isopropyl-4-thiazolyl)methyl]amino]carbonyl-l-valine and N-deacylvaline ritonavir is thiazol-5-ylmethyl(2S,3S,5S)-5-[(S)-2-amino-3-methylbutanamido]-3-hydroxy-1,6-diphenylhexan-2-ylcarbamate. 

b Thiazol-5-ylmethyl (2S,3S,5S)-5-acetamido-3-hydroxy-1,6-diphenylhexan-2-ylcarbamate. 

c Bis(thiazol-5-ylmethyl) (2S,3S,5S)-3-hydroxy-1,6-diphenylhexane-2,5-diyldicarbamate. 

d Thiazol-5-ylmethyl (2S,3S,5S)-3-hydroxy-5-[(S)-2-(3-{[2-(2-hydroxypropan-2-yl)thiazol-4-yl]methyl}-3-methylureido)-3- methylbutanamido]-1,6-diphenylhexan-2-ylcarbamate. 

e Thiazol-5-ylmethyl (2S,3S,5S)-3-hydroxy-5-[(S)-4-isopropyl-2,5-dioxoimidazolidin-1-yl]-1,6-diphenylhexan-2-ylcarbamate. f Thiazol-5-ylmethyl (2S,3S,5S)-5-[(S)-2-(3-{[2-(2-hydroperoxypropan-2-yl)thiazol-4-yl]methyl}-3-methylureido)-3-methylbutanamido]-3- hydroxy-1,6-diphenylhexan-2-ylcarbamate. 

g(4S,5S)-Thiazol-5-ylmethyl 4-benzyl-5-{(S)-2-[(S)-4-isopropyl-2,5-dioxoimidazolidin-1-yl]-3-phenylpropyl}-2-oxooxazolidine-3-carboxylate. h Thiazol-5-ylmethyl (2S,3S,5S)-5-[(S)-2-{3-[(2-ethylthiazol-4-yl)methyl]-3-methylureido}-3-methylbutanamido]-3-hydroxy-1,6-diphenylhexan 2-ylcarbamate. 

i BOC-aminoalcohol is thiazol-5-ylmethyl (2S,3S,5S)-(5-t-butoxycarbonylamino)-3-hydroxy-1,6-diphenylhexan-2-ylcarbamate and isobutoxycarbonyl aminoalcohol is thiazol-5-ylmethyl (2S,3S,5S)-(5-isobutoxycarbonylamino)-3-hydroxy-1,6-diphenylhexan-2-ylcarbamate. j (S)-N-[(S)-1-[(4S,5S)-4-Benzyl-2-oxooxazolidin-5-yl]-3-phenylpropan-2-yl]-2-{3-[(2-isopropylthiazol-4-yl)methyl]-3-methylureido}-3- methylbutanamide. 

k( S)-Isobutyl 2-{3-[(2-isopropylthiazol-4-yl)methyl]-3-methylureido}-3-methylbutanoate. 

l Thiazol-5-ylmethyl (2S,4S,5S)-4-hydroxy-5-[(S)-2-{3-[(2-isopropylthiazol-4-yl)methyl]-3-methylureido}-3-methylbutanamido]-1,6- diphenylhexan-2-ylcarbamate. 

m Thiazol-5-ylmethyl (2S,3R,5S)-3-hydroxy-5-[(S)-2-{3-[(2-isopropylthiazol-4-yl)methyl]-3-methylureido}-3-methylbutanamido]-1,6- diphenylhexan-2-ylcarbamate. 

n Bis(thiazol-5-ylmethyl) (2S,2'S,3S,3'S,5S,5'S)-5,5'-carbonylbis(azanediyl)bis(3-hydroxy-1,6-diphenylhexane-5,2-diyl)dicarbamate. 

o Thiazol-5-ylmethyl (2S,3R,5R)-3-hydroxy-5-[(S)-2-{3-[(2-isopropylthiazol-4-yl)methyl]-3-methylureido}-3-methylbutanamido]-1,6- diphenylhexan-2-ylcarbamate. 

p Thiazol-5-ylmethyl (2S,3S,5R)-3-hydroxy-5-[(S)-2-{3-[(2-isopropylthiazol-4-yl)methyl]-3-methylureido}-3-methylbutanamido]-1,6- diphenylhexan-2-ylcarbamate. 

q(3S,4S,6S,10S,13S,15S,16S)-Bis(thiazol-5-ylmethyl)-4,15-dihydroxy-10-isopropyl-8,11-dioxo-3,6,13,16-tetrabenzyl-2,7,9,12,17- pentaazaoctadecanedioate. 

r (2S,2'S)-N,N'-[(2S,3S,5S)-3-Hydroxy-1,6-diphenylhexane-2,5-diyl]bis(2-{3-[(2-isopropylthiazol-4-yl)methyl]-3-methylureido}-3- methylbutanamide). 

s(S)-[(5S,8S,10S,11S)-8,11-Dibenzyl-5-isopropyl-1-(2-isopropylthiazol-4-yl)-2-methyl-3,6,13-trioxo-15-(thiazol-5-yl)-14-oxa-2,4,7,12- tetraazapentadecan-10-yl] 2-{3-[(2-isopropylthiazol-4-yl)methyl]-3-methylureido}-3-methylbutanoate. 

5 SPECIFIC TESTS 

Water Determination 〈921〉, Method I: NMT 0.5%, determined on 0.500 g 

6 ADDITIONAL REQUIREMENTS 

Packaging and Storage: Preserve in tight, light-resistant containers. Store between 5° and 30°. 

USP Reference Standards 〈11〉 

USP Ritonavir RS 

USP Ritonavir Related Compounds Mixture RS