Introduction

Organic Volatile impurities (OVIs) ₁ are residual solvents that are used in and are produced during the synthesis of drug substances, or in excipients used in the production of drug formulations. Many of these residual solvents generally cannot be completely removed by standard manufacturing processes or techniques and are left behind, preferably at low levels. These impurities encounter during manufacture, storage of API, excipients and drug products. ICH ₂ has given limits for the presence of OVIs in APIs ₃ .In manufacturing drug substances, organic volatile impurities (OVI’s) arising from the final purification by recrystalization, and also from one or more steps of the synthetic process, can be retained in the end products. Very often these solvents, referred to as residual solvents, are carried to the pharmaceutical preparation concerned, making their determination mandatory ₄ . During the manufacturing process, certain types of formulations like Gel extrusion module tablet (GEM) is exposed to several organic volatile chemicals. Ethanol, isopropyl alcohol and acetone are commonly used during granulation process. Methanol may be present in certain grades of ethanol. Ethanol and acetone are also used in the preparation of the polymeric coating of GEM tablets ₅ .Isopropyl alcohol may be used in the crystallization of the active ingredient while ethyl acetate is a process solvent for the gel forming polymer 6.Low levels of these organic solvents are inevitably present in the GEM drug product even after drying process. OVIs, not only affect physiochemical properties of a drug, such as particle size, dissolution rate and stability, but also can present a serious potential health hazard. Methods for the quantification of OVI’s are reported by the United States Pharmacopoeia (USP) ₇ and European Pharmacopoeia (EP) ₈ , where the wide spread but relatively expensive automated head space gas chromatography (HS-GC) instrument is used. With these methods relative standard deviations RSDs as high as 15% are allowed ₇₈ . However the methods mentioned in the USP and EP are dedicated to the determination of residual solvents in drug substances, excipients or drug products. The content of residual organic solvents in pharmaceuticals is routinely measured by gas chromatography (GC). Routine GC applications include the analysis of samples of active pharmaceutical ingredients and their intermediates to comply with good laboratory practices (GLP) and good manufacturing practices (GMP), as well as in- process testing for residual solvents to optimize the drying process ₉ . Over the last decade, several GC methods to monitor residual solvents in pharmaceutical samples have been reported in the literature _1011121314 . Several of these GC methods tend to have long run times and are very specific for a limited number of OVI’s. There are several methods reported for the determination of residual solvents. Residual solvents are detected using Hewlett Packard (Agilent, Wilmington), MODEL 6890 over column a 60m X0.53mm RTX-1701 column with thickness 1.0µm with FID using carrier gas system of helium ₁₅ . The detection limit were determined as 3 ppm for methanol, 2 ppm for ethanol and isopropyl alcohol and 1 ppm for acetone and ethyl acetate An ICH Class 3 solvent, 1-propanol has been determined in propylgallate sample (80 µg/g ppm) with Agilent (Wilmington, DE, USA) GC, over 60m X 0.53mm id RTX column with carrier gas system of helium with FID. With the same system ethyl acetate has been determined in carbomer (0.155 µg/g ppm). A module drug powder soluble in water was chosen and residues of four solvents ethanol, cyclohexane triethylamine and pyridine were investigated ₁₆ . Ethanol was the purification/crystallization solvent, cyclohexane was used to denature ethanol, triethylamine was reactant and pyridine was the extracting solvent .The above solvents were determined in drug powder (product) with Varian 3800 CX system connected to Varian 8200 CX auto sampler for SPME over CP-select 624 CB column (Chrompack, Les Vils, France) 30mX 0.25mm id, thickness of 1.8µm with retention time 4.9 min, 9.6 min,11.5 min and 15 min for ethanol, cyclohexane, triethylamine and pyridine respectively using FID with helium as carrier gas system ₁₆ .The residual solvents testing in samples of drug substance intermediates were determined by using Rapid GC and Flash GC methods. Rapid GC was performed on Agilent (Alto, CA) modles 5890 series II and 6890 column ARTX 502.2, 1.4µm film of diphenyl/dimethyl polysiloxane stationary phase 30 m 0.25mm id, with FID, static headspace injection was performed via a model 7694 headspace sampler from Agilent technologies. A Flash GC method was derived from Thermo Orion application note with narrow bore column 10mX0.1mm id, with a 0.4 µm film thickness, 6% cyanopropyl, 94% polydimethyl siloxane [9]. In the reported method using rapid GC, sample A consisting of methanol (0.34%w/w), ethanol (2.64%w/w), and THF (0.21%W/W), sample B consisting of ethanol (0.04%w/w), sample C consisting of dichloromethane (0.05%w/w) and toluene (0.06%w/w), sample D consisting of toluene (0.03%w/w) and p-xylene (0.26%w/w) were determined accurately with good sensitivity . Samples were analysed with EZ Flash GC.

Weiyong Li ₁₇ reported the GC separation on Agilent DB-wax column with a dimension of 30m x 0.53mm and film thickness of 1µm. Helium was used as carrier gas with flow rate of 5ml/min with FID. The samples were injected with the Agilent 6890 series auto sampler in splitless mode. The detection limit (LOD) of the method for the mysilate esters was estimated from a chromatogram of a solution containing about 0.04µg/ml each of the esters, signal to noise ratio of 16, 25 & 29 for Methyl methane sulfonate (MMM), Ethyl methyl sulfonate (EMS) and Isopropyl methane sulfonate (IPMS), in pharmaceuticals where mesylate esters are known to be potent mutagenic, carcinogenic and teratogenic compounds LOQ was determined to be less than or equal to 5µg/g (5ppm) for MMS, EMS or IPMS based on the presion and accuracy data.

Costing C Camarasu ₁₈ developed headspace SPME method for the analysis of volatile polar residual solvents by GC-MS. The GC-ion trap mass spectrometer (GC-MS) was used with 30m x 0.25mm id. SPB-1 column coated with 1µm thickness (supleco park Bellefoute PA, USA) using helium as carrier gas system, the external electron ionization ion source was operated at an electron energy of 70ev and the filament emission was set at 200mA. SPME method has been developed and optimized for the polar residual solvent determination in pharmaceutical products. Five different polymers were investigated & the caboxen/polydimethylsiloxane was found to be the most sensitive for all compounds. Two head space SPME methods were developed and optimized one for the extraction from aqueous solutions and the other for the extraction from organic solvents N,N-dimethyl formamide (DMF) and dimethyl sulfoxide (DMSO). It was found that the addition of 100ml DMSO or DMF to 50 mg drug substance and slightly pressurizing the head space vial gave good results in terms of sensitivity & reproducibility. The detection limit were between 0.4 & 200 ng/ml of RSD on data were between 2 & 9 %. The headspace SPME from aqueous solution was found to be 10 times more sensitive than immersion SPME and head space SPME from organic solutions. Raquel Otero. etal ₁₉ have developed HS-GC method for the quantitative determination of residual solvents in a drug substance according to the European Pharmacopoeia method. A 6890 series Hewlett pack and GC system with a FID system (Waldbronn, Germany) and a 7496 HP headspace auto sampler equipped with a 1mL sample loop were used. An OVI-G43capillary column (30m × 0.53mm id) and 3µm film thickness (Supleco) was used. Solvent water-DMF (3:2) was selected to obtain good recoveries for ethanol, tetrahydrofuran. Same sample dilution sample was adopted to detect all class 1and 2 solvent at ICH recommended levels. By FID except 1, 1, 1- trichloroethane found to at 10 ppm instead of 500 ppm (ICH limit). Koji Urakami ₂₀ reported the influence of the matrix media used for the determination of residual solvents in pharmaceuticals by static headspace gas chromatography for water soluble samples. Water is the matrix medium of choice, for water insoluble samples, dimethyl sulfoxide (DMSO), N -dimethyl formamide (DMF), N,N-dimethyl acetamide (DMA), benzyl alcohol (BA), and 1,3-dimethyl-2- imidazolidine (DMI) are recommended by many authors ₂₁₂₂₂₃ . The author has also given some information about the formation of artifacts arising from the matrix medium heating the sample solution in a HS sampling apparatus or from ultrasonication of the sample solution used to increase sample solution. A Shimadzu GC-2010 gas chromatography equipped with FID and Perkin Elmer HS-40 headspace injector was used, with column 30x 0.53 mm id, fused silica capillary coated with 3.0µm film thickness of 6% cyanopropylphenyl 94% dimethyl polysiloxane (OVI-G43, Spleco Co.Ltd.Bellefonte,PA,USA) with heium as carrier gas system. Yunfei sha ₂₄ reported a head space solid-phase microextraction (HS-SPME) coupled to GC-MS for the determination of volatile compounds in a famous Chinese medicinal prescription, Xiao- Cheng –Qi –Tang. Some parameters affecting the extraction efficiency such as stirring, extraction temperature, fiber exposer time and desorption time were optimized. The results were obtained using a 100µm poly dimethyl siloxane (PDMS), during headspace extraction at 100 rpm for 20 min. Twenty seven compounds were identified in Xiao Cheng-Qi-Tang including some main compounds such as d-Limonene and linalol. Analysis was carried out on HP 6890 GC system, coupled with a HP MD 5973 quadrupole mass spectrometer, using HP-5MS capillary column (30m x 0.25mm id x 0.25µm flm thickness), with helium as the carrier gas system at flow rate of 1ml/min. Saeed Nojavan ₂₅ reported the relationship between residual solvents concentration and ampicillin trihydrate crystals stability. The amount of residual solvents determied by GC, x-ray powder diffraction (XRPD) and fourier transform Infrared spectroscopy (FT-IR) were used for characterization of solid state. The results have good relationship between concentration of methylene chloride (as a critical residual solvent) and degree of ampicillin trihydrate crystallinty with the increasing methylene chloride concentration in the sample. Analysis of the residual solvents was carried out on a Varian Modle 3600 GC, (Varian Iberica,Madrid,Spain) with FID using CP- Sil-5 capillary column (25m x 0.32mm id x 0.53µm film thickness), helium was the carrier gas system at a flow rate of 1ml/min ₂₅ . J. P. Guzowski ₂₆ has developed GC method for measuring hydroxylamine (HA) in a variety of sample matrix including pharmaceutical formulations. This article relies on converting HA into nitrous oxide (N₂O) which a single step reaction carried out directly in a heated headspace vail. The gaseous products were then analysed by headspace capillary GC, HP 5890, equipped with GC-Q megabore column (30m x0.53 mm id, P/N 1153432) with helium as carrier gas system. The electron capture detector (ECD) provided the best sensitivity (4 ppb). HA was efficiently recovered (98%) from a sample matrix that contained only the drug substance, however, a lower recovery was achieved (83%), when excipients were present. Mary- Annedep Barrio ₂₇ Carried out the simultaneous determination of formic acid and formaldehyde in pharmaceutical excipients using Agilent model 6890N GC equipped with a model 5973 N quadrupole mass selective detector (MSD) and Model 7694 headspace auto sampling unit with column Phenomenex ZB-WAX column (100% polyethylene glycol), 30m long with 0.32mm id and 0.5µm film thickness, with helium as the carrier gas system. The amount of formic acid and formaldehyde present in various excipients were lactose, formicacid (1.0ppm) and formaldehyde (< 0.2ppm), powdered cellouse, formic acid (2.9ppm) and formaldehyde (0.4ppm), mannitol, formic acid (0.0ppm) and formaldehyde (0.0ppm), microcrystalline cellulose 50µm, formic acid(9.3ppm) and formaldehyde (<0.2ppm), starch 1500 formic acid (3.0ppm) and formaldehyde (<0.2ppm) and magnesium sterate, formic acid (2.1ppm) and formaldehyde (0.9ppm). In present article GC 17 A Shimadzu was used with Flame Ionization detector (FID). The separation was carried out on BP 624 column (30m X 0.53mm i.d. X 0.25µm coating thickness), with nitrogen as carrier gas in the split mode by direct injection method. The method described was simple, sensitive, rugged, reliable and reproducible for the quantitation of acetone, ethyl acetate, isopropyl alcohol, methanol, tetrahydrofuran and toluene at residual level from intermediates and APIs.

Experimental Method

Instruments and Materials

Preparation of standard

Preparation of Sample

Gas chromatographic conditions

Method Validation

Results and Discussion

Development of method

Gas chromatographic method for the determination of residual solvents in API was developed. The column used was BP624 capillary column, flow rate 3.2 mLmin ^-1 , linear velocity 22 cmsec ^-1 and column pressure 14kpa with total flow of 116mLmin ^-1 in the split mode. In the prescribed method all six solvents eluted within 16 min (Fig. 1). The retention time for solvents is given in Table 1. The APIs have shown presence of various residual solvents in different concentrations (Table 2).

Figure 1

Table 1: Report of standard solvent chromatograms

Figure 2

Figure 1: Standard chromatograms of six solvents

Figure 3

Table 2: Determination of residual solvents in APIs

Validation of method

Specificity

Precision

Precision of the method and system are expressed in terms of standard deviation and relative standard deviation. For the precision of method and system the %RSD for six solvents complies with the acceptance criteria of less than 2% (Table 3.1). Hence the method and system is said to be precise.

Figure 4

Table 3.1: Precision data

Linearity

All six solvents showed broad range of linearity (Fig.2). The correlation coefficient calculated for all six solvents lies in the acceptance criteria of more than 0.99 (Table 3.2).

Figure 5

Figure 2: Standard calibration curves for six solvents

Limit of Detection (LOD) and Limit of Quantitation (LOQ)

The LOD and LOQ were calculated by statistical methods. For the instrumental method LOD is determined as the lowest amount for detection and LOQ is the lowest amount to quantifiable by the detector. LOD and sLOQ were determined by statistical formula.

LOD = 3.3SD/slop LOQ = 10SD/slope, Where, SD is standard deviation

The values for the Limit of Detection and Limit of Quantification for acetone, ethyl acetate, isopropyl alcohol, methanol, tetrahydrofuran and toluene are mentioned in Table 3.

Figure 6

Table 3.2: Linearity, LOD and LOQ data

Ruggedness

The ruggedness of method was established by performing the experiments on same chromatographic system with the same column by two analysts on a different day. The assay results of ruggedness studies were in the range of 90-105% v/v (Table 3.3). Additionally, good separation between the peaks of standard was achieved, which indicated that the method was selective for all components under the test.

Figure 7

Table 3.3: Ruggedness data

Conclusion

This study presents a simple and validated Gas Chromatographic method for estimation of residual solvents in intermediates and APIs. The method developed is specific, accurate, precise and rugged. The amount of organic volatile impurities present in the intermediates and APIs were found to be within the ICH limits.

Acknowledgement

Correspondence to

S. B. Puranik Department of Quality Assurance, Al Ameen College of Pharmacy, Opp. Lalbagh Main Gate, Hosur Road, Bangalore-560 027. Karnataka (India) Phone no. 011-91-9980231925 Fax no. 011-91+ 80 22225834/22278464 Email: sangpur@rediffmail.com