Analysis of USP Method <467> Residual Solvents on the Agilent 8890 GC System
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1 Application Note Residual Solvent Analysis of USP Method <6> Residual Solvents on the Agilent 889 GC System Author Lukas Wieder, Jie Pan, and Rebecca Veeneman Agilent Technologies, Inc. 8 Centerville Road Wilmington DE 988 Abstract This Application Note highlights the use of the Agilent 889 GC and the Agilent J&W DB-Select 6 UI for 6 and Agilent J&W HP-INNOWax columns in the detection and confirmation of <6> residual solvents. The system meets all specifications required in USP Method <6>, and demonstrates excellent repeatability across several injections.
2 Introduction Class and class residual solvents must be monitored and regulated, and the method for the analysis of these solvents involves three procedures: Procedure A: Initial identification and limit test using a G3 phase column (in this case, Agilent J&W DB-Select 6 UI for 6) Procedure B: If above limit in procedure A, perform a confirmation of peak identity and secondary limit test using a G6 phase column (in this case, Agilent J&W HP-INNOWax) Procedure C: If above limit in procedures A and B, perform a quantification using whichever column provided fewer coelutions This Application Note analyzed residual solvents listed in USP Method <6> with the Agilent 889 GC. The J&W DB-Select 6 UI for 6 and J&W HP-INNOWax columns were used in this analysis, and configured with dual flame ionization detectors (FIDs). Therefore, procedures A and B could be run simultaneously with one injection. Experimental S/S Inlet 889 GC Consumables Table. Consumables and part numbers. Consumable Description Vials -ml Clear crimp top headspace vials (p/n 9 8) Septa Advanced Green nonstick inlet septa (p/n 83 9-) Splitter Ferrules Deactivated fused silica Inert tee for capillary flow technology (p/n G38-66) Short graphite for. to.3 mm columns, /pk (p/n 8-883) UltiMetal Plus flexible metal, for.3 mm fused silica tubing, /pk (p/n G388-) Inlet liner mm, Splitless, straight, deactivated (p/n 8 888) Headspace transfer line/pre-cft column HSS transfer line Inert tee FID CH Column J&W DB-Select 6 UI for 6 Column J&W HP-INNOWAX FID CH 69A headspace sampler Figure. Experimental setup using a dual-column, dual-fid configuration for the analysis of USP <6> residual solvents. Deactivated Fused Silica, 3 m. mm id.3 mm od (p/n 6--3) Equipment An 889 GC was configured with a split/splitless inlet (SSL) and dual FIDs, and sampling was performed using an Agilent 69A headspace sampler. An inert tee was used to split the flow equally between the two columns, with both columns leading directly to FIDs. Figure shows the complete configuration. Chemicals and reagents Dimethyl sulfoxide (99.9 %) and water (HPLC grade) were purchased from Sigma-Aldrich. Column Column J&W DB-Select 6 UI for 6, 3 m.3 mm,.8 µm (p/n 3-33UI) J&W HP-INNOWax, 3 m.3 mm,. µm (p/n 99N-3I)
3 Sample preparation Sample preparation for the residual solvent samples was performed according to the USP <6> protocol. Three stock solutions of residual solvents in DMSO were used: Residual Solvent Revised Method <6> Class (p/n 9-9) Residual Solvent Revised Method <6> Class A (p/n 9-9) Residual Solvent Revised Method <6> Class B (p/n 9-9) The sample preparation procedures for each of the three classes are listed below: Class solvents. One milliliter of stock solution vial plus 9 ml of DMSO diluted to ml with water. One milliliter from step diluted to ml with water 3. Ten milliliters from step diluted to ml with water. One milliliter from step 3 with ml of water into headspace vial Class A solvents. One milliliter of stock solution vial, diluted to ml with water. One milliliter from step with ml of water into headspace vial Experimental parameters Table. System parameters for the analysis of residual solvents. GC system parameter 889 GC Carrier gas Helium constant flow mode, ml/min on column Inlet type Split/splitless Inlet temperature C Mode Split mode, split ratio : Oven C (hold min) to 8 C at 8 C/min (hold 3 min) Column flow ml/min in constant flow mode, column flow controlled by column FID (both channels) C Air ml/min H Makeup (N ) Headspace parameter Sample loop 3 ml/min ml/min 69A headspace sampler ml Oven temperature 8 C Loop temperature 8 C Transfer line temperature C Vial equilibration time minutes Injection duration. minutes Vial size ml Vial shaking On, level ( shakes/min) Vial fill mode Default: flow to pressure Vial fill pressure psi Loop ramp rate psi/min Final loop pressure psi Loop equilibration time. minutes Software Agilent OpenLab CDS Version. Class B solvents. One milliliter of stock solution vial, diluted to ml with water. One milliliter from step with ml of water into headspace vials 3
4 Results and discussion In addition to showing clear chromatography on both columns for each class of solvents and consistent results across many runs, there are requirements described in USP <6> the analysis must meet. Class solvents Figures to illustrate the analysis of classes, A, and B residual solvent mixes on the J&W DB-Select 6 UI for 6 and J&W HP-INNOWax GC columns. Analysis of class solvents meets the signal-to-noise ratio (S/N) and resolution requirements on both the J&W DB-Select 6 UI for 6 and J&W HP-INNOWax columns. The area and retention time repeatability measurements (RSD%) were evaluated on a set of headspace vials. Tables 3 to list the RSD% obtained on the J&W DB-Select 6 UI for 6 and J&W HP INNOWax columns for class, A, and B residual solvent mixes. The resulting RSD% values were less than. %, indicating high repeatability and stability of the column, the 69A headspace sampler, and the 889 GC/FID system Compound RT.,-Dichloroethane.8.,,-Trichloroethane.9 3. Carbon tetrachloride.8. Benzene 8.6.,-Dichloroethane 8.9 Figure. Chromatogram of the USP residual solvents class standard solution resolved on a J&W DB-Select 6 UI for 6 GC column.
5 ,3 Compound RT.,-Dichloroethane.8.,,-Trichloroethane.9 3. Carbon tetrachloride.9. Benzene 3.3.,-Dichloroethane Figure 3. Chromatogram of the USP residual solvents class standard solution resolved on a J&W HP-INNOWax column. Table 3. Repeatability (n = ) for class residual solvents obtained on the J&W DB-Select 6 UI for 6 and J&W HP-INNOWax columns. Compound Area RSD (%) on J&W DB Select 6 UI for 6 RT RSD (%) on J&W DB Select 6 UI for 6 Area RSD (%) on J&W HP INNOWax RT RSD (%) on J&W HP INNOWax,-Dichloroethane ,,-Trichloroethane Carbon tetrachloride.9.6 Coelutes with,,-trichloroethane Coelutes with,,-trichloroethane Benzene ,-Dichloroethane
6 Class A solvents Compound RT 3.6 3,. Methanol Acetonitrile Methylene chloride trans-,-dichloroethene.38. cis-,-dichloroethene Tetrahydrofuran Cyclohexane Methylcyclohexane 9. 9.,-Dioxane 9... Toluene.83.. Chlorobenzene.6.8. Ethylbenzene. 3. m-xylene.8.6. p-xylene.8.. o-xylene Figure. Chromatogram of the USP residual solvents class A standard solution resolved on a J&W DB-Select 6 UI for 6 GC column. 3 Compound RT 3.. Methanol 3.. Acetonitrile Methylene chloride 3.8. trans-,-dichloroethene.9.6. cis-,-dichloroethene Tetrahydrofuran 3.8. Cyclohexane.. 8. Methylcyclohexane.. 9.,-Dioxane Toluene Chlorobenzene Ethylbenzene m-xylene.98.. p-xylene o-xylene , Figure. Chromatogram of the USP residual solvents class A standard solution resolved on a J&W HP-INNOWax column. 6
7 Table. Repeatability (n = ) for class A residual solvents obtained on the J&W DB-Select 6 UI for 6 and J&W HP-INNOWax columns. Compound Area RSD (%) on J&W DB Select 6 UI for 6 RT RSD (%) on J&W DB Select 6 UI for 6 Area RSD (%) on J&W HP INNOWax RT RSD (%) on J&W HP INNOWax Methanol Acetonitrile Methylene chloride trans-,-dichloroethene cis-,-dichloroethene Tetrahydrofuran.3.9 Coelutes with methylene chloride Coelutes with methylene chloride Cyclohexane..9.. Methylcyclohexane..9..6,-Dioxane Toluene Chlorobenzene....3 Ethylbenzene.... m-xylene p-xylene Coelutes with m-xylene Coelutes with m-xylene..6 o-xylene....3 Class B solvents Compound RT. 8. n-hexane Nitromethane Chloroform.39.8.,-Dimethoxyethane 8... Trichloroethane Pyridine Hexanone Tetralin Figure 6. Chromatogram of the USP residual solvents class B standard solution resolved on a J&W DB-Select 6 UI for 6 GC column. 6
8 . Compound RT n-hexane.8. Nitromethane Chloroform. 3..,-Dimethoxyethane Trichloroethane Pyridine Hexanone Tetralin Figure. Chromatogram of the USP residual solvents class B standard solution resolved on a J&W HP-INNOWax GC column. 6 Conclusion The 889 GC system, equipped with a 69A headspace sampler and inert tee, provides an excellent method for separating, identifying, and quantifying all of the relevant residual solvents outlined by USP <6>. Beyond the expected coelutions, the peaks in all three classes are well resolved from each other, showing sufficient S/Ns, and can be quantified repeatably. Reference Table. Repeatability (n = ) for class B residual solvents obtained on the J&W DB-Select 6 UI for 6 and J&W HP-INNOWax columns. Compound Area RSD (%) on J&W DB-Select 6 UI for 6 RT RSD (%) on J&W DB-Select 6 UI for 6 Area RSD (%) on J&W HP-INNOWax RT RSD (%) on J&W HP-INNOWax n-hexane...9. Nitromethane Chloroform..8..,-Dimethoxyethane Trichloroethylene Pyridine Hexanone Tetralin USP 3-NF, General Chapter USP <6> Organic volatile impurities, United States Pharmacopeia. Pharmacopoeia Convention Inc., Rockville, MD, USA. This information is subject to change without notice. Agilent Technologies, Inc. 9 Printed in the USA, February, 9 99-EN
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