Revised USP 467 Residual Solvent Method

Satisfy New Method Requirements with Restek Columns and Standards

By Rick Lake, Pharmaceutical Innovations Chemist
  • Overview of the new USP 30/NF 25 procedure, 2nd supplement & interim revision, effective July 2008.
  • New reference standards - stock USP mixes, custom preparations.
  • Improve system suitability pass rates by choosing the correct liner size.

Background

Organic Volatile Impurities (OVIs), which are commonly referred to as residual solvents, are trace level chemical residues in drug substances and drug products that are byproducts of manufacturing, or that form during packaging and storage. It is the responsibility of the drug manufacturer to ensure that these residues are removed, or are present only in limited concentrations. The International Conference on Harmonization (ICH) publishes a guideline (Q3C) listing the acceptable amounts of solvent residues that can be present. In the ICH guideline, residual solvents are summarized by class, according to their toxicity. Class 1 compounds are carcinogenic compounds that pose a risk to both the consumer and the environment. The use of these solvents is to be avoided, but if they are used, they must be tightly controlled. Class 2 compounds are nongenotoxic animal carcinogens, and concentrations of these compounds should be limited. Chromatographic analysis is needed for both the Class 1 and Class 2 residual solvents. Class 3 compounds, on the other hand, have low toxic potential and may be assayed by nonspecific techniques.

The USP general chapter <467> Residual Solvents is a widely used compendial method used for identifying and quantifying residual solvents when there is no information available on what solvents are likely to be present. In an attempt to better mirror the ICH guidelines, the USP is proposing a more comprehensive methodology in residual solvent testing—the current USP30 NF25. This revised method is more comprehensive, as it increases the number of residual solvents to be routinely tested to 32. This is a much longer analyte list than previously tested and represents the sum of Class 1 and 2 residual solvents. The actual number of analytes may be more if xylenes, ethyl benzene and cis/trans 1,2 dichloroethylene are differentiated or if circumstances require the quantification of specific Class 3 residual solvents. The test method itself is also more extensive and divided into three distinct procedures (A, B and C) for identification, confirmation, and quantification.

Initially set to become effective July 1, 2007, the implementation of the current version of USP <467>: Residual Solvents has been delayed until July 1, 2008. Until that time, the Other Analytical Procedures section of the previous version will be retained. However, looking forward to the implementation of the revised method, this application will comply with the procedure and criteria set forth in the USP30 NF25, second supplement (effective December 1, 2007) and the interim revision announcement.

Overview of Method

The revised analysis of residual solvents under USP <467> consists of a static headspace extraction coupled with a gas chromatographic separation and flame ionization detection (HS-GC/FID). ICH Class 1 and 2 compounds need chromatographic determination and therefore make up the residual solvent reference standards used in this analysis. The routine analyte list is comprised of the Class 1 and Class 2 compounds and is separated into 3 test mixes — Class 1 Mixture, Class 2 Mixture A and Class 2 Mixture B. Class 2 Mixture C contains higher boiling solvents not readily detectable by headspace analysis and therefore has been omitted in the second supplement of USP30 NF25 or the interim revision announcement. These solvents are to be assayed by other appropriately validated procedures.

USP <467> is divided into two separate sections based upon sample solubility: water soluble and water insoluble articles. Under the updated methodology, the diluent used in standard and sample preparations differs for the water-soluble and water-insoluble articles; water is used for water-soluble articles, whereas dimethylformamide or dimethylsulfoxide is used for water-insoluble articles. However, the methodology for both water-soluble and water-insoluble articles is very similar. The test method for both sections consists of three procedures (A, B and C) that are designed to identify, confirm and then finally quantify residual solvents found to present in drug substances and products.

Autosamplers are widely used in headspace analysis and are also included in the USP <467> monograph. We have purposely performed this application using two different types of headspace autosamplers—a pressurized loop system and a heated syringe. Procedure A was performed using a Tekmar HT3 pressured loop autosampler to demonstrate implementing analyses using a transfer line. Procedure B was performed using an Overbrook Scientific HT200H to demonstrate implementing analyses using a syringe injection.

Procedure A - Identification

Procedure A is the first step in the identification process and is performed on a G43 (Rtx®-1301 or 624) column, as a means of determining if any residual solvents are present in the sample at detectable levels. First, Class 1 standard and system suitability solutions and a Class 2 Mix A standard solutions are assayed under the given operating conditions to determine suitability of the chromatographic system. All peaks in the Class 1 system suitability solution must have a signal-to-noise ratio not less than 3, the Class 1 standard solution must have a 1,1,1-trichloroethane response greater than 5, and in the Class 2 Mixture A solution the resolution of acetonitrile and dichloromethane must not be less than 1. When system suitability has been achieved, the test solutions are then assayed along with the Class 1 and Class 2 Mixtures A and B standard solutions. In the event that a peak is determined in the sample that matches a retention time and has a response greater than that of a corresponding reference material, the analyst then proceeds to Procedure B for verification of the analyte. An exemption is made for 1,1,1-trichloroethane, where a response greater than 150 times the peak response denotes an amount above the percent daily exposure limit. Figures 1 through 3 illustrate the analysis of residual solvent mixes Class 1 and Class 2 A and B by Procedure A. The resolution between acetonitrile and dichloromethane is easily achieved using an Rtx®-1301 (or 624) column and a 1mm split liner, coupled to a transfer line injection.


Figure 1  USP Residual Solvent Class 1 standard solution on an Rtx®-624 (G43) column.

Peaks
1.1,1-Dichloroethene
2.1,1,1-Trichloroethane
Peaks
3.Carbon Tetrachloride
4.Benzene
5.1,2-Dichloroethane
Residual Solvents Class 1 on Rtx®-624
GC_PH00909
ColumnRtx®-624, 30 m, 0.32 mm ID, 1.80 µm (cat.# 10970)
SampleResidual Solvents - Class 1 (cat.# 36279)
Injectionheadspace-loop split (split ratio 5:1)
Liner:Siltek® 1mm Split liner (cat.# 20972-214.1)
Headspace-Loop
Inj. Port Temp.:140 °C
Instrument:Tekmar HT3
Inj. Time:1.00 min
Transfer Line Temp.:105 °C
Valve Oven Temp.:105 °C
Sample Temp.:80 °C
Sample Equil. Time:45 min
Vial Pressure:10 psi
Pressurize Time:0.5 min
Loop Pressure:5 psi
Loop Fill Time:2.00 min
Oven
Oven Temp.:40 °C (hold 20 min) to 240 °C at 10 °C/min (hold 20 min)
Carrier GasHe, constant flow
Flow Rate:2.16 mL/min
Linear Velocity:35.3 cm/sec
DetectorFID @ 240 °C
NotesSample: USP <467> Class 1 standard solution
(cat.# 36279) in 20mL headspace vial

Figure 2  USP Residual Solvent Class 2 Mixture A standard solution on an Rtx®-624 (G43) column.

Peaks
1.Methanol
2.Acetonitrile
3.Dichloromethane
4.trans-1,2-Dichloroethene
5.cis-1,2-Dichloroethene
6.Tetrahydrofuran
7.Cyclohexane
Peaks
8.Methylcyclohexane
9.1,4-Dioxane
10.Toluene
11.Chlorobenzene
12.Ethyl benzene
13.m-Xylene/ p-Xylene
14.o-Xylene
Residual Solvents Class 2 Mixture A on Rtx®-624
GC_PH00910
ColumnRtx®-624, 30 m, 0.32 mm ID, 1.80 µm (cat.# 10970)
SampleResidual Solvents Class 2 - Mix A (cat.# 36271)
in 20 mL headspace vial
Injectionheadspace-loop split (split ratio 5:1)
Liner:Siltek 1mm Split (cat.# 20972-214.1)
Headspace-Loop
Inj. Port Temp.:140 °C
Instrument:Tekmar HT3
Inj. Time:1.00 min
Transfer Line Temp.:105 °C
Valve Oven Temp.:105 °C
Sample Temp.:80 °C
Sample Equil. Time:45 min
Vial Pressure:10 psi
Pressurize Time:0.5 min
Loop Pressure:5 psi
Loop Fill Time:2.00 min
Oven
Oven Temp.:40 °C (hold 20 min) to 240 °C at 10 °C/min (hold 20 min)
Carrier GasHe, constant flow
Flow Rate:2.16 mL/min
Linear Velocity:35.3 cm/sec
DetectorFID @ 240 °C

Figure 3  USP Residual Solvent Class 2 Mixture B standard solution on an Rtx®-624 (G43) column.

Peaks
1.Hexane
2.Nitromethane
3.Chloroform
4.1,2-Dimethoxyethane
5.Trichloroethylene
6.Pyridine
7.2-Hexanone
8.Tetralin
Residual Solvents Class 2 Mixture B on Rtx®-624
GC_PH00911
ColumnRtx®-624, 30 m, 0.32 mm ID, 1.80 µm (cat.# 10970)
SampleResidual Solvents Class 2 - Mix B (cat.# 36280)
in 20 mL headspace vial
Injectionheadspace-loop split (split ratio 5:1)
Liner:Siltek 1mm Split (cat.# 20972-214.1)
Headspace-Loop
Inj. Port Temp.:140 °C
Instrument:Tekmar HT3
Inj. Time:1.00 min
Transfer Line Temp.:105 °C
Valve Oven Temp.:105 °C
Sample Temp.:80 °C
Sample Equil. Time:45 min
Vial Pressure:10 psi
Pressurize Time:0.5 min
Loop Pressure:5 psi
Loop Fill Time:2.00 min
Oven
Oven Temp.:40 °C (hold 20 min) to 240 °C at 10 °C/min (hold 20 min)
Carrier GasHe, constant flow
Flow Rate:2.16 mL/min
Linear Velocity:35.3 cm/sec
DetectorFID @ 240 °C
NotesSample: USP <467> Class 2 Mixture B standard solution
(cat.# 36280) in 20mL headspace vial

Procedure B - Confirmation

Once a residual solvent is identified and found to be above the percent daily exposure limit, Procedure B is then performed to confirm analyte identity. A G16 capillary column (Stabilwax®) is used here as a confirmation column because it yields an alternate selectivity when compared to that of a G43 column. The same residual solvent mixes used in Procedure A, both standard and system suitability preparations, are again analyzed on this column. The system suitability requirements differ here in that the Class 1 standard solution must have a benzene response greater than 5 and the resolution of acetonitrile and cis-dichloroethene must not be less than 1 in the Class 2 Mixture A solution. If the analyte identified in Procedure A again matches the retention time and exceeds the peak response of the reference materials, again with the same exception to 1,1,1-trichloroethane, the analyst must now quantify the analyte using Procedure C. Figures 4 through 6 illustrate the analysis of residual solvent mixes Class 1, Class 2A and B on a Stabilwax® column. Again, the system suitability on a Stabilwax® column, used in conjunction with a 2 mm liner and syringe headspace injection, is easily achieved.


Figure 4  USP Residual Solvent Class 1 standard solution on a Stabilwax® (G16) column.

Peaks
1.1,1-Dichloroethene
2.1,1,1-Trichloroethane
Peaks
3.Carbon tetrachloride
4.Benzene
5.1,2-Dichloroethane
Residual Solvents (Class 1) on Stabilwax® (G16)
GC_PH00951
ColumnStabilwax®, 30 m, 0.32 mm ID, 0.25 µm (cat.# 10624)
SampleResidual solvents - class 1 (cat.# 36279)
in 20 mL headspace vial (cat.# 24685)
Diluent:Water
Injection
Inj. Vol.:1.0 µL headspace-syringe split (split ratio 5:1)
Liner:2 mm splitless (cat.# 20712)
Inj. Temp.:140 °C
Headspace-Syringe
Instrument:Overbrook Scientific HT200H
Syringe Temp.:100 °C
Sample Temp.:80 °C
Sample Equil. Time:45 min
Inj. Speed:8
Inj. Dwell:5 sec.
Oven
Oven Temp.:50 °C (hold 20 min) to 165 °C at 6 °C/min (hold 20 min)
Carrier GasHe, constant flow
Flow Rate:2.15 mL/min
Linear Velocity:35.2 cm/sec
DetectorFID @ 250 °C

Figure 5  USP Residual Solvent Class 2 Mixture A standard solution on a Stabilwax® (G16) column.

Peaks
1.Cyclohexane
2.Methylcyclohexane
3.trans-1,2-Dichloroethene
4.Tetrahydrofuran
5.Methanol
6.Dichloromethane
7.cis-1,2-Dichloroethene
Peaks
8.Acetonitrile
9.Toluene
10.1,4-Dioxane
11.Ethyl benzene
12.p-Xylene
13.m-Xylene
14.o-Xylene
15.Chlorobenzene
Residual Solvents (Class 2 - Mixture A) on Stabilwax® (G16)
GC_PH00952
ColumnStabilwax®, 30 m, 0.32 mm ID, 0.25 µm (cat.# 10624)
SampleResidual solvents class 2 - mix A (cat.# 36271)
in 20 mL headspace vial (cat.# 24685)
Diluent:Water
Injection
Inj. Vol.:1,000 µL headspace-syringe split (split ratio 5:1)
Liner:2 mm splitless (cat.# 20712)
Inj. Temp.:140 °C
Headspace-Syringe
Instrument:Overbrook Scientific HT200H
Syringe Temp.:100 °C
Sample Temp.:80 °C
Sample Equil. Time:45 min
Inj. Speed:8
Inj. Dwell:5 sec.
Oven
Oven Temp.:50 °C (hold 20 min) to 165 °C at 6 °C/min (hold 20 min)
Carrier GasHe, constant flow
Flow Rate:2.15 mL/min
Linear Velocity:35.2 cm/sec
DetectorFID @ 250 °C

Figure 6  USP Residual Solvent Class 2 Mixture B standard solution on a Stabilwax® (G16) column.

Peaks
1.Hexane
2.1,2-Dimethoxyethane
3.Trichloroethylene
4.Chloroform
5.2-Hexanone
6.Nitromethane
7.Pyridine
8.Tetralin
Residual Solvents (Class 2 - Mixture B) on Stabilwax® (G16)
GC_PH00953
ColumnStabilwax®, 30 m, 0.32 mm ID, 0.25 µm (cat.# 10624)
SampleResidual solvents class 2 - mix B (cat.# 36272)
in 20 mL headspace vial (cat.# 24685)
Diluent:Water
Injection
Inj. Vol.:1,000 µL headspace-syringe split (split ratio 5:1)
Liner:2 mm splitless (cat.# 20712)
Inj. Temp.:140 °C
Headspace-Syringe
Instrument:Overbrook Scientific HT200H
Syringe Temp.:100 °C
Sample Temp.:80 °C
Sample Equil. Time:45 min
Inj. Speed:8
Inj. Dwell: 5 sec.
Oven
Oven Temp.:50 °C (hold 20 min) to 165 °C at 6 °C/min (hold 20 min)
Carrier GasHe, constant flow
Flow Rate:2.15 mL/min
Linear Velocity:35.2 cm/sec
DetectorFID @ 250 °C

Procedure C - Quantification

Lastly, when a residual solvent is identified and verified in both Procedures A and B, the analyst then performs Procedure C to quantify the analyte. This is done by analyzing the sample against the specific, individual reference material for the analyte identified. Individual standards are prepared by diluting the analyte in solution to a concentration of 1/20 of the concentration limit given in the method. Following the procedure given and the instrument conditions in either Procedure A or B, depending upon which procedure provides the most definitive results, a quantifiable result is produced.

Conclusion

Implementing the revised method for US <467> can be difficult if you do not optimize correctly. Although the second supplement contains a change that allows for modifications to the split ratio, liner and column choices are still important. Both the Rtx®-1301 (624) and the Stabilwax® columns can easily pass system suitability criteria, and the use of smaller bore liners (see Tech Tip) can increase peak efficiency. Restek can supply all your USP <467> materials and can help you optimize your testing within the constraints of the method.

Thank You!

Instruments provided courtesy of Overbrook Scientific™ and Teledyne Tekmar.

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