A Generic Method for the Analysis of Residual Solvents inPharmaceuticals Using StaticHeadspace-GC-FID/MS

AbstractThe determination of residual solvents in pharmaceuticals is one of the most impor-

tant gas chromatography (GC) applications in quality assurance/quality control

(QA/QC) in the pharmaceutical industry. Sample introduction is normally done using

static headspace (SHS). In routine QC, GC with flame ionization detection (FID) is pre-

ferred, while mass spectrometry (MS) can be used for screening and identification. In

this application note, a retention-time locked GC-MS/FID method is presented that

allows the analysis of more than 50 solvents in a single run. The method was opti-

mized to allow identification and quantification of all three International Conference

on Harmonisation (ICH) classes of solutes at relevant concentration levels.

AuthorsKarine Jacq, Frank David, and

Pat Sandra

Research Institute for Chromatography,

Pres. Kennedypark 26, B-8500 Kortrijk,

Belgium

Matthew S. Klee

Agilent Technologies, Inc.

2850 Centerville Road

Wilmington, DE 19808

USA

Application Note

Pharmaceuticals

2

IntroductionThe determination of residual solvents (RS), formerly calledOrganic Volatile Impurities (OVI), in pharmaceutical productsis probably the most important application of gas chromato-graphy (GC) in pharmaceutical quality control. Recently, meth-ods described in U.S. and European Pharmacopoeia havebeen reviewed, updated, and harmonized according toInternational Conference on Harmonisation (ICH) guidelineQ3C (R3) [1].

In pharmaceutical manufacturing, approximately 60 differentsolvents are in typical use. This set of solvents covers arather large range of boiling points and polarities. Accordingto the ICH guideline, these solvents are divided into threeclasses. Class 1 includes benzene, carbon tetrachloride, 1,1-dichloroethane, 1,2-dichloroethylene, and 1,1,1-trichloro-ethane; these solvents are toxic and their use should beavoided. Class 2 solvents are less toxic, but their use shouldalso be limited. Class 1 and Class 2 solvents are preferablybeing replaced by Class 3 solvents, which have low toxicpotential to humans. Taking into account their relative toxici-ty, these solvents should be monitored in pharmaceuticalproducts, including drug substances (or active pharmaceuticalingredients [API]) and drug products (formulations) at variouslevels, ranging from 2 ppm (2 µg/g drug substance) for ben-zene (Class 1) to 0.05 % (w/w = 5,000 ppm) for Class 3 sol-vents. Consequently, the analytical method(s) used to monitorthese residual solvents in pharmaceutical products needsalso to cover this range.

For the analysis of residual solvents, gas chromatography incombination with flame ionization detection (GC-FID) is nor-mally used. Sample preparation and introduction is done bystatic headspace [2]. In this way, the (mostly) volatile sol-vents are introduced selectively and the analytical system(inlet, column, and detector) is not contaminated by the(mostly) nonvolatile drug substance or drug product. For theseparation, a thick film, medium polar column (for example,G43) is selected. Quantification is done versus an externalstandard.

Excellent quantitative data, including low limits of detection(LODs) high repeatability and excellent linearity wereobtained using the Agilent G1888 static headspace sampler incombination with an Agilent 7890 GC [3]. However, since nocolumn can guarantee a unique retention time for a given sol-vent, confirmation analysis by GC-FID on a capillary columncoated with a different stationary phase (for example, G16) isperformed. More recently, GC-MS has been successfully usedfor confirmation/identification purposes [4,5].

In this application note, a system configuration and operationconditions are described that allow the analysis of 56 sol-vents in a single run. Some solvents listed in the ICH guide-line are not volatile (enough) and not amenable to SHS-GCanalysis. Examples are formic acid, acetic acid, dimethyl sul-foxide (often used as a method solvent), formamide, ethylene

glycol, and sulfolane. The analysis of these impurities shouldbe performed using other methods; their analysis is not dis-cussed here.

The presented retention-time locked method, however, can beconsidered as generic, since it covers most solvents and canbe used both for identification (by MS) and for quantification(by FID and/or MS) of residual solvents across a wide con-centration range.

Experimental Sample PreparationFor the analysis of residual solvents in pharmaceutical prod-ucts, the drug substance or drug product is typically dissolvedin a low-volatility (high-boiling) solvent such as dimethyl sul-foxide (DMSO), dimethyl acetamide (DMAC), or 1,3-dimethyl-2-imidazolidinone (DMI). For water-soluble drug substances,dissolution in water can also be used. In this work, 100 mgdrug substance was dissolved in 2 mL DMSO or DMSO/water(1:1). Recently dedicated "GC headspace" -grade solventswere made available from Sigma-Aldrich (NV/SA Bornem,Belgium). DMSO, suitable for GC-HS (cat. no. 51779) wasused in this work.

Solvent standard solutions were prepared in DMSO at 15 µg/mL concentration for Class 1 solvents and at 600 µg/mL concentration for Class 2 and Class 3 solvents.From these stock solutions, aliquots of 1 to 100 µL wereadded in a standard 20 mL HS vial filled with 2 mL DMSO/water (1:1, v/v). The concentrations of standards are alwaysexpressed in microgram per gram of drug product or drug sub-stance. So, the concentration of these calibration solutionsranges from 0.15 to 15 ppm (µg/g drug) for Class 1 and from 6to 600 ppm for Class 2 and Class 3 solvents, if 100 mg productis weighed in the HS vial. (The actual concentration of stan-dards in the vial, expressed in µg per mL solvent, is 20 timeslower.)

Instrumental ConditionsThe samples were analyzed using the SHS-GC-FID/MS con-figuration presented in Figure 1. Static headspace was per-formed using a G1888 HS autosampler. The transfer line ofthe headspace sampler is coupled to a standard split/split-less inlet. Separation was done on a DB-1301 column. Thecolumn effluent is split using a purged splitter Capillary FlowTechnology device to FID and MS (5975 MSD). The vial pres-sure is regulated by an AUX EPC channel. The purge at thesplitter is also regulated by the AUX EPC (second channel). A63 cm × 0.1 mm id deactivated fused silica capillary was usedto connect the splitter to the MSD; a 40 cm × 0.1 mm id capil-lary was used to connect the splitter to the FID. Flows in bothcapillaries are approximately 1.4 mL/min and the retentiontime is also similar (small offset between FID and MS retention times). The analytical and selected ion monitoring(SIM) parameters are summarized in Table 1 and Table 2,respectively.

3

Results and DiscussionColumn SelectionMany different columns can be used for the analysis of resid-ual solvents after static headspace extraction. Typically, a col-umn coated with a thick film of a medium polar stationaryphase is selected. A classical column for residual solventanalysis is a 30 m × 0.53 mm id coated with 1 to 3 µm DB-624 (β = 44). On this column, good separation is obtainedin about 30 minutes of analysis time [3].

Table 1. Analytical Parameters

SHS (G1888 Agilent HS autosampler)Loop size: 1 mL Vial pressure: 14 psig (96.5 kPa)Headspace oven: 80 °CLoop temperature: 120 °CTransfer line temperature: 120 °CEquilibration time: 10 min, high shakePressurization: 0.15 minVent (loop fill): 0.5 minEquilibration time: 0.1 minInject: 0.5 min

GC (7890A)Inlet: Split/splitless, split 1/10, headspace liner

(P/N 5183-4709)Inlet temperature: 250 °CSplit ratio: 1:10Carrier gas: HeliumInlet pressure: 160 kPa *AUX pressure (at splitter): 60 kPaColumn: DB-1301 20 m × 0.18 mm × 2 µm (J & W)Oven: 40 °C (5 min) – 5 °C/min – 80 °C – 10 °C/min

– 200 °C (2 min)FID: 300 °C, 40 mL/min H2, 400 mL/min air

MS (5975 C Inert XL MSD)Transfer line: 300 °CMode: Simultaneous scan/SIMSCAN range: 29 – 250 m/zSIM: See Table 2* Retention time locking was applied. The column head pressure was adjusted to elutetoluene at 16.160 min.

Table 2. SIM Parameters

Group Start time Ions1 0.00 31, 302 3.50 31, 43, 45, 593 4.60 31, 43, 45, 58, 59, 61, 74, 964 5.55 41, 43, 845 6.50 57, 61, 73, 966 7.25 41, 57, 867 7.90 31, 42, 598 8.70 30, 43, 61, 72, 969 9.65 42, 45, 47, 59, 72, 8310 10.30 47, 56, 61, 84, 97, 11711 11.10 43, 45, 49, 61, 62, 74, 76, 78, 9012 12.02 43, 7113 12.70 31, 56, 95, 13014 13.40 55, 8315 13.90 43, 58, 61, 8816 14.50 31, 59, 7117 15.60 41, 43, 52, 55, 58, 73, 79, 9118 16.75 42, 55, 7019 17.30 43, 58, 10020 17.66 43, 56, 7321 18.20 77, 91, 106, 11222 20.50 44, 78, 87, 105, 108, 12023 21.30 44, 98, 9924 26.00 91, 104, 132

Figure 1. SHS-GC-FID/MS system configuration.

In this work, a narrow-bore, thick-film column with equivalentstationary phase, DB-1301 (equivalent to G43), was selected.The lower phase ratio (β = 22) results in better resolution forthe first eluting (but frequently detected) solvents, such asmethanol, ethanol, diethylether, acetone, isopropanol, and acetonitrile.

It is not possible to find a stationary phase that is able to sep-arate all 60 solvents in approximately 30 minutes. In mostpractical cases, however, only three to five solvents need tobe determined. The described method allows the analysis of56 solvents and eventually the same column can also be usedfor the analysis of ethylene oxide. For this reason, columnselection and separation conditions can be considered asgeneric.

4

The separation obtained by SHS-GC-FID/MS for a solvent testmixture in DMSO/water is shown in Figure 2. The concentra-tion of the solutes was 16 ppm for the five Class 1 solventsand 560 ppm for the Class 2 and Class 3 solvents. With oneinjection, three data files are obtained. The upper trace showsthe total ion chromatogram (TIC) obtained by MSD in scanmode, the middle trace shows the corresponding SIM data,and the lower chromatogram is the FID trace. Toluene elutesat 16.16 minutes in all three chromatograms and the offset in retention time between FID and MS is less than 0.03 minute or 2 seconds for all solutes.

A detailed view of the separation is shown in Figure 3, show-ing three elution windows of the FID trace. All solutes can bedetected and are labeled on the chromatograms except fortetrachloromethane (trace at 10.93 minutes), 2-methoxyethanol (co-elutes with benzene, 1,2-dimethoxyethane, and 1,2-dichloroethane), 2-ethoxyethanol (trace at 14.96 minutes),DMAC (co-elutes with cumene), and 1-methyl-2-pyrrolidone(trace at 25.19 minutes).

ValidationIn a previous application note [3] it was demonstrated thatthe quantitative data obtained using a G1888 SHS – 7890A GCcombination resulted in equal or better quantitative resultsthan a G1888 – 6890 GC combination. Electronic pressurecontrol of carrier gas and vial pressure results in excellentrepeatability and linearity. An optional pneumatic controlmodule can be used to control the vent pressure by backpressure regulation (BPR), resulting in increased sensitivity(by a factor of 2 to 4) and smaller relative standard deviations(RSDs) (reduced by a factor of 2). In this study, good quantita-tive data were obtained even though the BPR approach wasnot used. Using BPR would provide higher performance forthose seeking it.

Limits of detection, linearity, and repeatability were evaluatedwith the different modes of detection. Repeatability waschecked at 3 ppm level (µg/g, equivalent to 0.15 µg/mL invial) for Class 1 solvents and at 100 ppm for the others (n = 6). Five-point calibration curves (plus a blank) were mea-sured between 0.15 and 15 ppm for Class 1 solvents, andbetween 6 and 600 ppm for most others. For some solutesgiving low response, linearity was tested in the 500 to 5,000 ppm range. For coeluting compounds, the experimentswere performed with single compound solutions in order toallow peak integration with FID. The results are summarizedin Table 3.

Column 4 in Table 3 indicates possible coelution of targetcompounds. Most solutes are chromatographically resolvedand can thus be quantified by either FID or MS when presentin the same sample. In some cases, coelution is observed(labeled with C). In these cases, quantification is still possibleby MS after ion extraction or by using selected ion monitoring(SIM) mode.

In columns 5 through 8, the limit of detection (LOD) for thethree detection modes, determined from the lowest calibra-tion level at S/N = 3, are compared to the ICH limits. In mostcases (43 of the 56 analytes), the LODs were well below theICH limits for all three detection modes. Those instanceswherein the LOD was above the ICH limit are highlighted inTable 3 in boldface. For the Class 1 solvents, it is clear thatMS in SIM mode is preferred. The benefit of using SIM isillustrated in Figure 4a for some Class 1 solvents. At 10.6 min-utes, 1,1,1-trichloroethane coelutes with cyclohexane, asseen in the FID trace. However, both solutes can be accurate-ly measured with extracted ion chromatograms using m/e =97 for 1,1,1-trichloroethane and m/e = 56 for cyclohexane.The same can be observed in Figure 4b for the overlappedpeaks of benzene, 1,2-dimethoxyethane and 1,2-dichloro-ethane at 11.5 minutes in the FID trace and the extracted ionpeaks of m/e = 78 for benzene, m/e 45 for 1,2-dimethoxy-ethane, and m/e = 62 for 1,2-dichloroethane (2-methoxy-ethanol is not detected at this level).

Using MS, all compounds, except 2-methoxyethanol can bedetected at LOD < ICH limit. Some of the other more polarsolutes (2-ethoxyethanol, DMF, DMAC, and 1-methyl pyrroli-done) also give a low response in FID, whereas MS detectionlimits are satisfactory. As reported before [3], back-pressureregulation of the vent pressure is expected to help decreasethe LOD by a factor of two, so this an important considerationif you are using FID for quantitation. In addition, increasingsample size and/or injection volume (headspace sample loopsize) would also help if you are doing quantitation with FID.

The repeatability of the SHS-GC-FID/MS method is excellent.RSDs were mostly below 5 percent, both for GC-FID and GC-MS (SIM and SCAN). The average RSDs were 4.4 percent forSCAN, 3.8 percent for SIM, and 3.0 percent for FID. Again, thevalues were higher for some more polar solutes. Finally, goodlinearity was obtained for most compounds. Except for 2-methoxyethanol, 2-ethoxyethanol, DMF, DMAC, and 1-methyl-2-pyrrolidone (compounds with higher LODs), linear-ity was excellent with the three types of detection (R² > 0.99).

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Figure 2. A 56-component solvent mixture analyzed by SHS-GC-FID/MS.

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Figure 3. Detailed FID chromatogram (16 ppm Class 1 and 560 ppm Class 2 and Class 3 solvents).

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Solvent InfluenceThe influence of the solvent used to dissolve the drug sub-stance or drug product was also evaluated. In general, equallygood quantitative data in terms of linearity and repeatabilityare obtained when using water, DMSO, DMAC, DMI, or mix-tures thereof. The LOD (and slope of the calibration curve),however, depends on the solvent used. In general, for apolarsolvents in the most polar matrix (water), the lowest LODswill be obtained. For the (apolar) Class 1 solvents, the LODcan be up to 10 lower (more sensitive) if static headspace isperformed in DMSO/water versus pure DMSO.

Matrix InfluenceThe presence of a relatively high concentration of drug sub-stance in the sample solution (100 mg/2 mL) might influencethe absolute response of the solutes. To evaluate that poten-

7

Table 3. Validation Results for 56 Solutes

tial with this method, some SHS-GC-FID/MS analyses wereperformed on solutions of 100 mg drug substance (prometh-azine) in the solvent (2 mL DMSO/H2O). A blank (not spiked)and two spiking levels (for example, 3 and 8 ppm for Class 1solvents, and 110 and 280 ppm for Class 2 and 3 solvents)were analyzed in triplicate.

Both repeatability (RSDs) and linearity (R²) were comparablewith the data obtained without active pharmaceutical ingredi-ent. In general, the responses for the spiked samples werebetween 80 and 105 percent of the response obtained with-out active pharmaceutical compound (API). This recovery iswell within the limits generally accepted for trace impurity analysis.

Methanol 2 2.77 No 3000 14.2 0.29 132 8.4 3.5 6.0 0.997 0.996 0.999

n-pentane 3 3.88 No 5000 3.3 0.09 6.2 3.7 2.8 1.1 0.994 0.994 0.998

Ethanol 3 4.11 No 5000 18.2 0.80 132 3.6 3.0 8.3 0.995 0.995 0.996

Ethyl ether 3 4.35 No 5000 1.4 0.06 6.0 3.5 3.5 1.3 0.996 0.995 1.000

1,1-dichloroethene 1 4.86 No 8 1.9 0.02 7.4 15.9 4.0 7.3 0.992 0.996 0.990

Acetone 3 5.01 No 5000 4.7 0.13 20.1 4.9 4.0 2.4 0.994 0.995 1.000

2-propanol 3 5.37 C 5000 17.0 0.12 230 4.0 3.3 2.2 0.997 0.997 1.000

Ethyl formate 3 5.38 C 5000 14.1 0.33 27 3.1 3.3 0.8 0.998 0.997 1.000

Acetonitrile 2 5.71 No 410 11.1 0.46 73 4.1 3.5 4.2 0.995 0.996 0.999

Methyl acetate 3 5.84 No 5000 2.8 0.16 40 2.5 3.8 2.8 0.997 0.997 1.000

Dichloromethane 2 6.06 No 600 1.7 0.06 24 4.6 4.0 2.4 0.997 0.996 0.999

Z-1,2-dichloroethene 2 6.74 C 1870 0.9 0.01 9.0 4.1 4.0 1.3 0.996 0.996 1.000

t-butyl methyl ether 3 6.73 C 5000 2.4 0.02 7.5 3.2 3.3 1.4 0.996 0.996 1.000

n-hexane 2 7.42 No 290 1.2 0.03 4.4 2.5 3.2 0.9 0.994 0.994 0.999

1-propanol 3 8.11 No 5000 23 0.45 126 6.6 3.6 9.1 0.995 0.995 0.998

Nitromethane 2 9.13 No 50 104 1.0 214 7.8 1.5 11.3 0.995 0.997 0.992

E -1,2-dichloroethene 2 9.24 Partial 1870 3.9 0.09 16 4.0 4.0 2.6 0.997 0.996 1.000

2-butanone 3 9.33 Partial 5000 11.1 0.32 40 3.3 3.4 3.1 0.997 0.997 0.999

Ethyl acetate 3 9.50 No 5000 12.4 0.5 19.1 3.4 4.3 3.8 0.997 0.997 0.999

2-butanol 3 9.82 No 5000 18.4 1.6 64 7.0 3.9 2.0 0.993 0.995 0.999

RSD r2

Class 1: __ 3 ppm Class 1: 0.15–15 ppmICH RT Co- LOD (ppm, rel. 100 mg API) Class 2 and 3: __ 100 ppm Class 2 and 3: 6–600 ppm

Compound class (min) elution1 ICH SCAN SIM FID SCAN SIM FID SCAN SIM FID

8

THF 3 9.96 No 5000 3.5 0.22 20 3.3 3.6 3.0 0.998 0.997 1.000

chloroform 2 10.07 Partial 60 1.6 0.04 34 3.0 3.7 3.8 0.998 0.997 0.995

1,1,1-trichloroethane 1 10.51 C 1500 2.1 0.03 18.8 4.7 3.3 6.0 0.997 0.997 0.993

Cyclohexane 2 10.62 C 3880 1.9 0.03 2.90 3.3 3.5 1.0 0.995 0.996 1.000

Tetrachloromethane 1 10.93 No 4 0.71 0.04 45 5.7 3.3 6.0 ² 0.998 0.997 0.994³

Isobutyl alcohol 3 11.32 No 5000 18 1.7 64 4.9 3.2 7.3 0.998 0.996 0.998

Benzene 1 11.48 C 2 0.70 0.02 4.8 6.2 3.5 2.4 0.994 0.997 1.000

2-methoxyethanol 2 11.48 C 50 835 248 2650 20 ² 9.5 ² 16 ² 1 point 0.988³ 0.868³

1,2-dimethoxyethane 2 11.46 C 100 13.2 < 3.5 65 4.5 3.2 5.5 0.995 0.995 0.999

1,2-dichloroethane 1 11.54 C 5 8.2 0.70 30 2.3 ² 4.0 1.6 ² 0.999³ 0.997 0.999³

Isopropyl acetate 3 11.74 No 5000 1.4 0.07 13.5 3.2 4.3 1.3 0.996 0.997 1.000

n-heptane 3 12.18 No 5000 1.3 0.06 3.5 3.3 3.6 1.8 0.995 0.995 0.999

1-butanol 3 13.05 C 5000 21.1 1.1 178 6.9 5.1 1.1 0.994 0.996 1.000

Trichloroethene 2 13.15 C 80 1.2 0.03 16 3.3 4.0 1.6 0.998 0.996 0.999

Methylcyclohexane 2 13.61 No 1180 0.6 0.02 3.8 3.7 3.8 1.1 0.996 0.996 1.000

1,4-dioxane 2 14.08 No 380 30 0.41 141 3.5 3.5 4.4 0.992 0.997 0.994

Propyl acetate 3 14.22 No 5000 1.3 0.20 16.2 3.6 3.9 1.8 0.997 0.997 1.000

2-ethoxyethanol 2 14.96 No 160 576 115 1000 5.0 ² 14.1 3.0 ² 0.977³ 0.956 0.989³

4-methyl-2-pentanone 3 15.80 No 5000 5.8 0.16 11.9 3.0 3.3 0.9 0.996 0.997 1.000

Isoamyl alcohol 3 15.94 No 5000 73 1.6 76 5.6 4.3 3.6 0.995 0.996 0.999

Pyridine 2 15.95 No 200 46 3.1 63 3.7 3.3 4.3 0.993 0.995 0.999

Toluene 2 16.16 No 890 0.4 0.02 3.6 4.2 3.9 1.1 0.997 0.997 1.000

Isobutyl acetate 3 16.50 No 5000 5.4 0.44 10.1 3.4 4.1 0.9 0.997 0.997 0.999

1-pentanol 3 17.01 No 5000 19.3 0.94 84 4.1 3.9 4.5 0.995 0.995 0.998

2-hexanone 2 17.55 No 50 1.8 0.10 15.0 2.1 3.4 1.0 0.997 0.997 1.000

n-butyl acetate 3 17.80 No 5000 0.9 0.08 12.0 4.2 3.6 0.8 0.997 0.997 0.999

DMF 2 18.59 No 880 962 192 2885 12.2 ² 10.9 ² 17.8 ² 0.970³ 0.977³ 0.937³

Chlorobenzene 2 19.03 No 360 0.7 0.03 8.5 3.5 3.9 1.3 0.996 0.996 0.998

Xylene 2 19.23 No 2170 2.2 0.15 29 4.6 3.8 2.2 0.996 0.997 0.999

Xylene 2 19.45 No 2170 0.5 0.03 5.9 3.8 3.8 1.2 0.997 0.997 0.999

Xylene 2 20.22 No 2170 2.4 0.16 25 3.7 3.8 2.7 0.997 0.996 0.999

DMAC 2 20.92 C 1090 154 18.8 3200 20 ² 17.3 ² 19.5 ² 0.914³ 0.921³ 0.935³

Cumene 3 20.92 C 5000 0.2 0.03 < 1 4.0 3.7 1.8 0.997 0.997 0.999

Anisole 3 21.07 No 5000 2.7 0.13 < 1 2.7 3.3 1.5 0.995 0.997 0.998

1-methyl-2-pyrrolidone 2 25.19 No 4840 6150 117 6150 ND² 12.1 ² ND² 1 point 0.972³ 1 point

Tetralin 2 26.72 No 100 1.7 0.12 12.5 4.2 3.5 1.3 0.995 0.992 0.997

ND = Not detected1 C = Coelution, but resolved by MS; Partial = partial overlap2 Calculated at 2,500 ppm3 Calculated in the 500 to 5,000 ppm range

RSD r2

Class 1: __ 3 ppm Class 1: 0.15–15 ppmICH RT Co- LOD (ppm, rel. 100 mg API) Class 2 and 3: __ 100 ppm Class 2 and 3: 6–600 ppm

Compound class (min) elution1 ICH SCAN SIM FID SCAN SIM FID SCAN SIM FID

Table 3. Validation Results for 56 Solutes (continued)

9

ConclusionsMore than 50 residual solvents can be determined in pharma-ceutical products in a single run using a static headspace –GC-FID/MS configuration. Quantification can be performedroutinely by FID for most target compounds, while MS isespecially suited for trace level determination of Class 1 sol-vents and for identification of unknowns. Mass spectrometryalso excels for determination of coeluting peaks through useof extracted ion or SIM ion chromatograms, thereby eliminat-ing the need for additional analyses on dissimilar columns.The quantitative data, including repeatability, linearity, andLOD are excellent, meeting or exceeding ICH guidelines.

Application

The method was applied to a number of available drug sub-stances. In samples of penicillin V and levamisol(tetramisole), traces of residual solvent were detected. As anillustration, the FID chromatograms for the two samples (foreach analysis, 100 mg sample was dissolved in 2 mLDMSO/water, 1:1) are shown in Figures 5 and 6. In the peni-cillin sample (Figure 5), a trace amount of n-butyl acetate wasdetected. The concentration, measured by external calibra-tion, was 52 ppm – well below the ICH limit (5,000 ppm). Inthe levamisol sample (Figure 6), a trace amount of toluenewas detected. The concentration was 66 ppm, well below the890 ppm ICH limit. It is interesting to note that a trace amountof dimethyl sulfide was detected in both chromatograms.DMS is an impurity in the DMSO solvent used to dissolve thesamples. In addition, an "unknown" was detected in lev-amisol. Through MS spectral library searching, the peak wasidentified as 2-chloropropane. This solvent is not included inthe ICH solvent list. However, the ability to unequivocallyidentify this unknown in the sample clearly demonstrates theadvantage of using parallel MS detection.

Solvent BackflushCapillary column backflushing is starting to be routinelyimplemented as a means of improving analysis cycle timesand data quality. The simplicity and benefits of implementingbackflush for residual solvents analysis were evaluated. Toreduce the analysis time, late-eluting solvents (DMSO andDMAC) can be backflushed by increasing the pressure at thecolumn outlet (AUX pressure controlling the purged splitter)and lowering the inlet pressure. This is demonstrated inFigures 7A through 7C. In a normal run of a solvent mixture,DMSO elutes at 20.8 minutes, and the standard run continuesto 200 °C to ensure that there is no carry-over of sample com-ponents into the next run. If no analytes of interest elute afterDMSO, backflushing can be used to reduce cycle times.Figure 7B shows the same analysis as in Figure 7A, only witha backflush initiated at 20 minutes (just before elution ofDMSO). The outlet pressure was increased from 60 to 200 kPa and the GC was held at 150 °C for 10 minutes. Nopeaks are observed after the backflush was initiated. Next, ablank run (without backflush, original method) was performed(Figure 7C). It clearly shows that DMSO solvent was totallyremoved. The ability to implement backflush was very simplebecause of the purged-split configuration of FID and MSD.Thermal stress on the column was certainly reduced and thecooldown time from 150 °C instead of the 200 °C original end-ing temperature was faster. Further improvement in cycletime can be achieved by reducing backflush time to the mini-mum time required to fully backflush DMSO.

Figure 4a. Comparison of overlapped peaks in FID and individual SIM ion chromatogramsfor 1,1,1-trichloroethane (m/e = 97) and cyclohexane (m/e = 56).

Figure 4b. Comparison of overlapped peaks in FID and individual SIM ion chromatogramsfor benzene (m/e = 78), dimethoxyethane (m/e = 45), and 1,2-dichloroethane (m/e = 62).

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Figure 5. Analysis of commercial penicillin sample. n-butyl acetate was determined to be present at 52 ppm – well below the ICH limit of 5,000 ppm.

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Figure 6. Analysis of levamisol sample. Toluene was determined to be present at 66 ppm – well below the ICH limit of 890 ppm.

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Figure 7A. Standards analysis without backflush.

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Figure 7B. Standards analysis with backflush at 20 minutes (just before DMSO elution).

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Agilent shall not be liable for errors contained herein orfor incidental or consequential damages in connectionwith the furnishing, performance, or use of this material.

Information, descriptions, and specifications in this publication are subject to change without notice.

© Agilent Technologies, Inc., 2008Printed in the USASeptember 18, 20085989-9726EN

References

1. ICH Harmonised Tripartite Guideline, Q3C(R3),http://www.ich.org/LOB/media/MEDIA423.pdf

2. R. L. Firor, “The Determination of Residual Solvents in Pharmaceuticals Usingthe Agilent G1888 Network Headspace Sampler,” Agilent Technologies publica-tion 5989-1263EN, 2004.

3. A. E. Gudat, R. L. Firor, and U. Bober, “Better Precision, Sensitivity, and HigherSample Throughput for the Analysis of Residual Solvents in PharmaceuticalsUsing the Agilent 7890A GC System with G1888 Headspace Sampler in DrugQuality Control,” Agilent Technologies publication 5989-6023EN, 2007.

4. R. L. Firor and A. E. Gudat, The Determination of Residual Solvents inPharmaceuticals Using the Agilent G1888 HS/6890GC/5975 inert MSDSystem,” Agilent Technologies publication 5989-3196EN, 2005.

5. A. E. Gudat and R. L. Firor, “The Determination of Extractables and Leachablesin Pharmaceutical Packaging Materials Using Headspace/GC/MS,” AgilentTechnologies publication 5989-5494EN, 2006.

For More InformationFor more information on our products and services, visit our Web site at www.agilent.com/chem.

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Figure 7C. Blank run after the backflush run demonstrates that the backflush completely removed DMSO.