High-speed gas chromatographic analysis of solvents in pharmaceuticals using solid phase microextraction

A simple, inexpensive and rapid analytical approach for the determination of organic volatile impurities in pharmaceutical drug substances is developed, where sample preparation step was conducted using solid phase microextraction (SPME), followed by a fast GC separation. With an extraction time between 3 and 5 min and separation of 13 solvents in less than 3 min employing fast temperature programming using resistively heated column, organic volatile impurities can be analyzed within a total analysis time of 6-9 min. Various SPME phases were evaluated towards sensitivity and selectivity for the extraction of 13 commonly found solvents in drug substances dissolved in dimethyl sulfoxide and water. A 2-cm Carboxen/polydimethyl siloxane/divinylbenzene (Carboxen/PDMS/DVB) phase and a 65-microm DVB/PDMS phase showed better sensitivity towards these solvents when extracted from organic and aqueous matrix in comparison with the sensitivity obtained with direct injection approach. Extraction parameters such as extraction time, extraction stir rate, etc. are discussed. %RSD of peak area of replicate extraction was between 2 and 10% when 100 microm PDMS was used for extracting solvents from aqueous matrix. When DVB/PDMS fiber was evaluated for precision, %RSD of peak area from replicate extractions of solvents from organic matrix was between 2 and 8%. One-hundred micrometer PDMS showed excellent linearity from 10 to 500 microg/ml for analytes extracted from water solutions. On the other hand, DVB/PDMS phase showed better linearity than Carboxen/PDMS/DVB fiber when it was used to extract analytes in the concentration range of 10-5000 microg/ml from organic matrix.     

Headspace SPME method development for the analysis of volatile polar residual solvents by GC-MS

A solid-phase microextraction (SPME) method has been developed and optimized for the polar residual solvent determination in pharmaceutical products. Five different polymer-coated fibers were investigated and the Carboxen/polydimethylsiloxane was found to be the most sensitive for all components. Two Headspace SPME methods were developed and optimized: one for the extraction from aqueous solutions, and the other for the extraction from organic solutions (N,N-dimethyl formamide (DMF) and dimethyl sulfoxide (DMSO). The optimum equilibration time for all components and all systems was 30 min. It was found that the sample headspace volume has an important effect on method sensitivity and precision. At low headspace volumes (less than one-third of the vial volume), sensitivity improves but at the same time, precision worsens. For 10 ml headspace vials, the optimum headspace volume was found to be 3 ml. The total volatile organic content in the sample also has an important effect on method sensitivity and precision. At low organic content, sensitivity increases but precision drops significantly. Over 0.5% volatile organic content in the sample, the system becomes unstable due to stationary phase swelling by the organic components, and also the sensitivity of the method is drastically reduced. The optimum range for total volatile organic content was found to be between 0.01 and 0.1%. The added Na2SO4 quantity increases the extraction yield. It was found that slightly pressurizing the headspace vial improves the sensitivity of the method by a factor of 2. For the organic system, it was found that the addition of 100 microl DMSO or DMF to 50 mg drug substance and slightly pressurizing the headspace vial gives good results in terms of sensitivity and reproducibility. The measured detection limits were between 0.4 and 200 ng/ml, and the relative standard deviation data were between 2 and 9%. The Headspace SPME from aqueous solutions was found to be ten times more sensitive than Immersion SPME and Headspace SPME from organic solutions.     

Rapid determination of tramadol in human plasma by headspace solid-phase microextraction and capillary gas chromatography-mass spectrometry

A simple, rapid and sensitive method for determination of tramadol in plasma samples was developed using headspace solid-phase microextraction (HS-SPME) and gas chromatography with mass spectrometry (GC-MS). The optimum conditions for the SPME procedure were: headspace extraction on a 65-microm polydimethylsiloxane/divinylbenzene (PDMS/DVB) fiber; 0.5 mL of plasma modified with 0.5 mL of sodium hydroxide (0.1 M); extraction temperature of 100 degrees C, with stirring at 2000 rpm for 30 min. The calibration curve showed linearity in the range of 1-400 ng mL(-1) with regression coefficient corresponding to 0.9986 and coefficient of the variation of the points of the calibration curve lower than 10%. The detection limit for tramadol in plasma was 0.2 ng mL(-1). The proposed method was successfully applied to determination of tramadol in human plasma samples from 10 healthy volunteers after a single oral administration.     

顶空气相色谱法测定羟乙基淀粉中有机溶剂残留量

目的 建立测定羟乙基淀粉中2种有机溶剂残留量的顶空气相色谱法。方法检测器为FID检测器,色谱柱为DB-FFAP毛细管柱（30m×0．53mm,1．5μm）,进样口温度为180℃,柱温为90℃,检测器温度为250℃。结果甲醇和环氧乙烷质量浓度线性范围分别为0．12～3．00mg／mL和0．0004～0．01mg／mL,可同时测定2组分。结论该方法操作简单,结果准确、可靠。     

Determination of delorazepam in urine by solid-phase microextraction coupled to high performance liquid chromatography

An SPME-HPLC-UV method for the determination of delorazepam, a representative benzodiazepine, in spiked human urine samples was developed for the first time. The performances of two commercially available fibers, a carbowax/templated resin (Carbowax/TPR-100) and a polydimethylsiloxane/divinylbenzene (PDMS/DVB), were compared, indicating the latter as the most suitable for urine samples analysis. All the aspects influencing adsorption (extraction time, pH, temperature, salt addition) and desorption (desorption and injection time, desorption solvent mixture composition) of the analyte on the fiber have been investigated. In particular, short extraction times were necessary to reach the equilibrium and very short desorption times were employed. The procedure required simple sample pre-treatment and was able to detect 5 ng/ml in spiked urine, regardless of the complexity of the matrix.     

Analysis of the volatile compounds in Ligusticum chuanxiong Hort. using HS-SPME–GC-MS

A headspace solid-phase microextraction (HS-SPME) method followed by gas chromatography–mass spectrometry (GC–MS) is described for the analysis of volatile compounds in the dry rhizome of Ligusticum chuanxiong Hort. Three types of SPME fibers including PDMS, PDMS–DVB and DVB–CAR–PDMS were investigated and the best extraction was achieved with the mixed fiber DVB–CAR–PDMS. Parameters for HS-SPME in terms of temperature and time, sample amount and particle size, and desorption time were also investigated. A polar capillary column was used for the chromatographic separation. As a result, 73 compounds were determined and identified by the HS-SPME–GC–MS method with at least 20 more compounds than those in the methods available. Comparison was made between HS-SPME–GC–MS and steam distillation (SD)-GC–MS methods. Using much less sample amount, shorter extraction time and simpler procedure, HS-SPME method can achieve similar results with those by SD. In conclusion, the present method is simple, rapid and effective and can be used for the analysis of volatile compounds in medicinal plants.     

Method development and validation for the GC-FID assay of p-cymene in tea tree oil formulation

This paper describes the development and validation of an isothermal gas chromatography-flame ionisation detection (GC-FID) method for the assay of pure tea tree oil. The chromatographic conditions of the method employ a 5% carbowax packed column (20 m x 0.25 mm), isothermal elution with hydrogen at a column flow of 36 ml/min, injector and detector temperature at 220 degrees C and oven temperature at 100 degrees C, and a 1.5 microl injection volume. Samples and standard were diluted in hexane. The calibration curve for p-cymene was linear (r2=0.9995) from 20 to 120% range of the analytical concentration of 100 microg/ml. The precision of this method was calculated as the relative standard deviation (R.S.D.) was 0.66% (n=6). The R.S.D. for intermediate precision study was 0.13 and recovery of the p-cymene ranged between 93.39 and 97.86%. The limits of detection and quantitation were determined to be 2.08 and 10.39 ng/ml, respectively.     

A comparison study of sample dissolution media in headspace analysis of organic volatile impurities in pharmaceuticals

Four sample dissolution media, N,N-dimethylacetamide (DMA), N,N-dimethyl-formamide (DMF), 1,3-dimethyl-2-imidazolidinone (DMI) and water were studied and compared in gas chromatographic headspace analysis of organic volatile impurities (OVIs) in drug substances. Emphasis was placed on fundamental parameters of headspace analysis such as maximum equilibrium temperature, precision, sensitivity and accuracy. The impacts of equilibrium time, temperature and salting out effect were determined and discussed in detail. It is found that the partition of an analyte in the sample solution influence not only the sensitivity but also the precision and accuracy of the analysis. Although the boiling point of DMI was much higher than those of DMF and DMA, they show the same maximum equilibrium temperature (110 °C). Salting out effect in aqueous sample solution diminishes the matrix effect of headspace analysis and increases the sensitivity of polar analytes. It is concluded that: (1) Water and DMI are the best hydrophilic and hydrophobic medium, respectively; (2) Anti-polarity effect is an approach to enhance the sensitivity; (3) Polar and high boiling point OVIs tend to involve problems of analysis precision, sensitivity and accuracy.     

顶空气相色谱法测定唑来膦酸中有机溶剂残留量

目的采用顶空气相色谱法测唑来膦酸中有机溶剂残留量。方法用DB-624毛细管柱（30m×0．32mm,1．8μm）,柱温为起始温度40℃,维持10min,再以每分钟35℃的速率升温至220℃,维持5min;进样口温度为250℃;氢火焰离子化检测器（FID）;检测器温度为280℃。结果6种溶剂的分离度良好,在所考察的质量浓度范围内线性关系良好,r=0．9999,平均回收率为96．9％-102．3％,检出限为7．86pg／mL-75．0ng／mL。结论所用方法简单、灵敏度高,可用于唑来膦酸中有机溶剂残留量的限度检查。     

顶空气相色谱法测定伊班膦酸钠中甲苯和氯苯的残留量

目的建立伊班膦酸钠原料药中有机溶剂甲苯和氯苯残留量检测的顶空气相色谱法。方法采用Agilent DB-624毛细管柱（30 m×0.53 mm,3μm）,程序升温方式,进样口温度180℃,检测器温度200℃,载气为N2,流速为5 mL/min,顶空瓶平衡温度为60℃,平衡时间为30 min。结果甲苯和氯苯分离良好,在所考察的质量浓度范围内具有良好的线性关系,r分别为0.999 3和0.999 4,平均回收率分别为90.96%和93.72%,精密度RSD均小于5%,最低检出质量浓度分别为0.35μg/mL和1.13μg/mL。3批样品中2种有机溶剂残留量均符合要求。结论所用方法简便快速、准确可靠,适于伊班膦酸钠原料药中有机溶剂残留量的检测。     

Determination of clenbuterol in human urine and serum by solid-phase microextraction coupled to liquid chromatography

A solid-phase microextraction (SPME)-LC-UV method for the determination of the beta-adrenergic drug clenbuterol in human urine and serum samples was developed for the first time using a polydimethylsiloxane/divinylbenzene (PDMS/DVB) coated fiber. The procedure required very simple sample pretreatments, isocratic elution, and provided highly selective extractions. All the aspects influencing fiber adsorption (extraction time, temperature, pH, salt addition) and desorption (desorption and injection time, desorption solvent mixture composition) of the analyte have been investigated. The linear ranges investigated in urine and serum were 10-500 and 5-500 ng/ml, respectively (that covers the typical clenbuterol concentration observed in biological fluids). Within-day and between-days R.S.D.% in urine ranged between 5.0-5.3 and 8.5-8.7, respectively, while in serum ranged between 5.5-5.9 and 8.7-9.1, respectively. Estimated LOD and LOQ were 9 and 32 ng/ml (spiked urine), respectively, and 5 and 24 ng/ml (spiked serum), respectively, well below the usual clenbuterol urinary and serum level.     

顶空气相色谱法测定福沙匹坦二甲葡胺中8种残留溶剂的含量

目的:建立顶空气相色谱法测定福沙匹坦二甲葡胺中甲醇、乙醇、丙酮、乙腈、甲基叔丁基醚、四氢呋喃、乙酸乙酯及甲苯残留量的方法。方法:采用顶空气相色谱法,色谱柱为DB-624毛细管柱(30 m×0.53 mm,3.0μm),载气为氮气,流速为2.0 ml·min^-1,进样口温度为200℃,FID检测器,检测器温度为250℃,程序升温:起始温度为45℃,维持5 min,再以20℃·min^-1的速率升温至200℃,维持3 min。顶空平衡温度为80℃,平衡时间为20 min。结果:8种残留溶剂分离度良好,在一定浓度范围内呈良好的线性关系(r≥0.9997),平均回收率在95%~105%范围内,检出限分别为0.19,0.31,0.06,0.13,0.03,0.05,0.16,0.12μg·ml^-1。结论:该方法灵敏度高,准确度好,适用于福沙匹坦二甲葡胺原料药中残留溶剂的测定。     

毛细管气相色谱法测定依诺昔酮原料药中6种有机溶剂残留量

目的:建立测定依诺昔酮原料药中6种有机溶剂(乙醇、丙酮、乙醚、二氯甲烷、乙酸乙酯与甲苯)残留量的方法。方法:采用毛细管气相色谱法。色谱柱为DB-1301毛细管柱;柱温采用程序升温;氢火焰离子化检测器,检测器温度为250℃;载气为高纯氮气;进样口温度为200℃;分流比为10∶1;顶空进样,加热温度为100℃,加热时间为40 min;进样瓶装量为2 ml。结果:乙醇、丙酮、乙醚、二氯甲烷、乙酸乙酯与甲苯在各自检测质量浓度范围内线性关系良好(r=0.999 6⁰.999 9);平均回收率为99.9%0.999 9);平均回收率为99.9%¹01.3%(RSD=0.76%101.3%(RSD=0.76%¹.89%,n=3);检测限分别为0.04、0.11、0.21、0.07、0.16、0.08 ng;3批样品中只检出乙醇。结论:建立的方法快速、灵敏、准确,可用于依诺昔酮原料药中有机溶剂残留量测定。     

顶空气相色谱法测定多烯磷脂酰胆碱注射液中的乙醇含量

目的：建立测定多烯磷脂酰胆碱注射液中助溶剂乙醇含量的方法。方法：采用顶空气相色谱法。色谱柱为DB—WAX毛细管柱,氢火焰离子化检测器,程序升温,进样口温度为200℃,检测器温度为220qC,顸空瓶平衡温度为65℃,平衡时间为7min,载气为氮气,分流比为5：1。以正丙醇为内标。结果：乙醇和正丙醇的分离度符合要求,乙醇检测质量浓度线性范围为0．1567～50．16mg／ml（r=0．9999,n=9）,平均回收率为100．07％,RSD：0．33％（n=6）,定量限为3．92pg／ml。结论：本方法操作简便,结果准确可靠,可用于多烯磷脂酰胆碱注射液中乙醇含量的控制。     

顶空气相色谱法测定枸橼酸喷托维林中有机溶剂残留量

目的建立测定枸橼酸喷托维林中有机溶剂残留量的顶空气相色谱法。方法采用FID检测器,色谱柱为HP-INNOWax毛细管柱（30m×0.53mm,1.0μm）,进样口温度180℃,柱温90℃,检测器温度250℃。结果在该试验条件下,乙醇和环氧乙烷质量浓度与峰面积线性关系良好,并可同时测定。结论顶空气相色谱法简单、准确、灵敏度高、重复性好,可用于该药物中有机溶剂残留量的测定。     

顶空气相色谱法测定注射用哌拉西林钠舒巴坦钠（4：1）残留溶剂含量

目的：用顶空气相色谱法测定注射用哌拉西林钠舒巴坦钠（4：1）中丙酮、二氯甲烷有机溶剂残留量。方法：利用顶空自动进样装置,外标法,在大口径弹性石英毛细柱上进行各组分分离,克服了进样量不准确的缺点,各组分间的分离度均符合要求。柱温：90℃;进样口温度：200℃;检测器温度300℃。顶空瓶平衡溫度80℃,平衡时间30分钟。结果：平均回收率：丙酮：100.6%二氯甲烷99.8%。结论：该方法利用顶空气相色谱法克服了进样量不准确,各组分分离度达不到要求的缺点,重现性好,精度高。     

Fast and fully automated analytical method for the screening of residues of aziridine and 2-chloroethylamine in pharmaceutical active principles

A simple, fast and fully automated method for the screening of aziridine (AZD) and 2-chloroethylamine (CEA) in active pharmaceutical ingredients (API) has been developed. The method is based on the in-fiber derivatization of the amines extracted from the sample headspace (previously dissolved or suspended in alkaline water) with 2,3,4,5,6-pentafluorobenzoyl chloride (PFBCl) previously adsorbed in the PDMS/DVB solid phase microextraction (SPME) fiber. The derivatives formed are further desorbed and analyzed in a gas chromatograph with negative ion chemical ionization mass spectrometry (GC-NCI-MS) using methane as reagent gas. The different operational parameters of the procedure have been optimized to get highest sensitivity. The validation of the method, however, revealed a poor repeatability, particularly evident in water-soluble APIs (RSD>20% for AZD). In spite of that, the low detection limits (1-3ngg(-1) for AZD and CEA), speed (44min total analysis time) and automation make that this method can be satisfactorily used as screening tool to accept or reject API batches attending to their volatile amine content and a critical specified value derived from the 1.5μg/day Threshold of Toxicological Concern (TTC) and maxima daily dosages. This was shown by analyzing seventy-five fluvoxamine maleate samples containing known levels of AZD and CEA (between 0.05 and 1.05μgg(-1)) in intermediate reproducibility conditions to get reliable estimations of precision and linearity. From these data, acceptance, rejection and non-conclusive areas of response are defined for both analytes at different confidence and replication levels using normal statistics. The method was satisfactorily applied to real fluvoxamine maleate samples.     

顶空气相色谱法测定舒萨林中残留溶剂含量

目的：用顶空气相色谱法测定舒萨林中乙醇、异丙醇、乙酸乙酯有机溶剂残留量。方法：利用顶空气相色谱法,在大口径弹性石英毛细柱上进行各组分分离,各组分间的分离度均符合要求。柱温：80℃;进样口温度：200℃;检测器温度250℃。顶空瓶平衡溫度80℃平衡时间30分钟。结果：平均回收率：乙醇100.8%异丙醇101.2%乙酸乙酯100.5%。结论：该方法利用顶空气相色谱法克服了各组分分离度达不到要求的缺点,重现性好,精度高。     

顶空气相色谱法测定盐酸克林霉素中的残留溶剂

目的：建立顶空毛细管气相色谱法测定盐酸克林霉素中有机溶剂残留量。方法：采用顶空气相色谱法,色谱柱为DB-624毛细管柱,载气为氮气,流速为2 ml·min^-1,分流比为1∶1。 FID检测器,检测器温度为300℃;进样口温度为200℃;柱温采取程序升温,起始温度40℃,保持5 min,以20℃·min^-1升温至200℃,持续10 min。顶空进样,平衡温度为80℃,平衡时间为30 min。以水为溶剂,外标法测定盐酸克林霉素中甲醇,乙醇,丙酮,三氯甲烷和N,N-二甲基甲酰胺的残留量。结果：在此色谱条件下,各有机溶剂均能得到有效分离,在所考察的浓度范围内线性关系良好,r=0.9991~0.9999;平均回收率为97.2%~101.2%,RSD为0.68%~1.98%（n=9）。结论：本试验建立的顶空气相色谱方法适合盐酸克林霉素原料中有机溶剂残留量的检测。