衍生气相色谱法测定甲磺酸伊马替尼中甲磺酸烷基酯类

目的建立衍生气相色谱法同时检测甲磺酸伊马替尼中的甲磺酸甲酯、甲磺酸乙酯、甲磺酸异丙酯三种甲磺酸烷基酯。方法采用NaI-H2O碘化衍生,利用ZB-WAX石英毛细管柱(30m×0.53mm,1.0μm)分离,氢火焰离子化(FID)检测器。结果方法在0.070~13.0μg/mL具有良好的线性(r0.999),检出限0.2ppm,回收率94.1%~111.3%。结论该方法可用于甲磺酸伊马替尼中甲磺酸甲酯、甲磺酸乙酯、甲磺酸异丙酯三种甲磺酸烷基酯成分的同时碘化衍生分离分析,方法简便,结果可靠、重现性好。     

LC-QQQ-MS/MS分析苯磺酸氨氯地平中痕量苯磺酸酯类基因毒性杂质

摘要：目的 建立LC-QQQ-MS/MS分析测定苯磺酸氨氯地平中痕量苯磺酸酯类基因毒性杂质苯磺酸甲酯、苯磺酸乙酯、苯磺酸丙酯和苯磺酸异丙酯的检测方法。方法 采用Agilent Zorbax SB-C18（3.0mm×100mm,1.8μm）色谱柱,以水（含 5mMol?L-1甲酸铵和0.1%甲酸）和甲醇（含5mMol?L-1甲酸铵和0.1%甲酸）为流动相,梯度洗脱,流速0.4 mL?min-1,电喷雾离子化（ESI）,正离子模式下MRM采集。结果 苯磺酸甲酯定量限为20 ng?mL-1,苯磺酸乙酯、苯磺酸丙酯和苯磺酸异丙酯定量限为1 ng·mL-1;方法精密度良好,保留时间和峰面积RSD%均小于5%（n=10）;苯磺酸甲酯在20~1000ng·mL-1,苯磺酸乙酯、苯磺酸丙酯和苯磺酸异丙酯在1~1000ng?mL-1的浓度范围内呈良好的线性关系,r≥0.998;方法准确度良好,平均加标回收率（n=10）分别为99.77%、100.19%、94.21%和92.43%。5个不同厂家生产的苯磺酸氨氯地平中苯磺酸甲酯的含量均小于LOQ（8μg?g-1）,苯磺酸乙酯、苯磺酸丙酯和苯磺酸异丙酯的含量均小于LOQ（0.4μg?g-1）。结论 该方法可用于苯磺酸氨氯地平中痕量基因毒性杂质苯磺酸甲酯、苯磺酸乙酯、苯磺酸丙酯和苯磺酸异丙酯含量的检测分析。     

GC-MS法同时测定盐酸决奈达隆中的3种甲磺酸酯

目的：建立同时测定盐酸决奈达隆中甲磺酸甲酯,甲磺酸乙酯,甲磺酸异丙酯含量的方法.方法：采用柱前衍生技术,利用GC-MS法,测定盐酸决奈达隆中三种甲磺酸酯的含量.结果：甲磺酸甲酯的检测限为0.003 75 ng·ml-1,线性范围为2.0 ～ 625.0ng·ml-1,r =0.999 4,甲磺酸乙酯的检测限为0.0375 ng·ml-1,线性范围为2.0 ～625.0 ng·ml-1,r=0.999 8,甲磺酸异丙酯的检测限为0.375 ng·ml-1,线性范围为2.0 ～625.0 ng·ml-1,r=0.999 8.回收率分别为99.89％,98.98％,100.36％,RSD分别为3.67％,2.82％,3.47％（n=9）.结论：该方法简便,准确,快速,适用于盐酸决奈达隆质量控制和检测.     

GC-MS法测定甲磺酸达比加群酯中的致突变杂质

目的 建立一种气相色谱-质谱联用法（GC-MS）同时测定甲磺酸达比加群酯中甲磺酸甲酯、甲磺酸乙酯和甲磺酸异丙酯的方法。方法 采用DB-624色谱柱（30 m×0.25 mm,1.4 μm）;程序升温：起始温度100 ℃,以15 ℃·min^-1的速率升温至200 ℃,再以25 ℃·min^-1的速率升温至240 ℃;进样口温度为220 ℃,载气为氦气,流速为1.0 mL·min^-1,进样量为2.0 μL。采集模式为选择离子监测（SIM）,离子源温度为230 ℃。通过内标法进行计算。结果 三种甲磺酸酯之间的分离度均大于2.0,甲磺酸甲酯、甲磺酸乙酯、甲磺酸异丙酯的线性回归方程分别为Y=0.0130X+0.0524（r=0.999 0）、Y=0.0249X+0.0633（r=0.999 3）、Y=0.0188X+0.0906（r=0.998 5）,均在5.0～120.0 ng·mL^-1浓度范围内线性关系良好。检出限为1.5 ng·mL^-1（0.15 ppm）,定量限为5 ng·mL^-1（0.5 ppm）。甲磺酸甲酯、甲磺酸乙酯和甲磺酸异丙酯的平均回收率分别为96.28%、97.91%和95.87%,RSD分别为2.83%、2.93%和1.73%。结论 本方法简便、快速、准确、专属性强、灵敏度高,适用于甲磺酸达比加群酯中三种甲磺酸酯的检测。     

顶空气相色谱-质谱法测定布南色林原料药中遗传毒性杂质

目的：建立顶空气相色谱-质谱法测定布南色林原料药中甲磺酸甲酯、甲磺酸乙酯和甲磺酸异丙酯3种遗传毒性杂质含量的方法。方法：采用顶空气相色谱-质谱法，以甲磺酸丁酯为内标，按内标标准曲线法进行甲磺酸甲酯、甲磺酸乙酯和甲磺酸异丙酯的含量测定。色谱条件：DB-WAX毛细管色谱柱（30 m×0.25 mm×0.25 μm）；程序升温，初始柱温为40℃，维持3 min，升温速率为30℃·min^-1，终止温度150℃，保持2 min；进样口温度为110℃；载气（He）流速为0.6 mL·min^-1；进样量为1 mL；进样方式为分流进样，分流比为20：1。质谱条件：电子轰击离子源（EI），扫描方式为选择性离子检测；离子源温度为200℃；接口温度为150℃；电子能量为70 eV；溶剂延迟1 min。结果：3种杂质成分之间的分离度均大于2.0；甲磺酸甲酯、甲磺酸乙酯、甲磺酸异丙酯检测质量浓度线性范围均为0.025~3.0 μg·mL^-1（r ≥ 0.998 5）；精密度、稳定性、重复性试验的RSD<5%；加样回收率分别为93.40%~101.40%（RSD为3.2%，n=9）、92.80%~99.70%（RSD为2.5%，n=9）和96.30%~100.75%（RSD为1.6%，n=9）。结论：该方法简便、准确、灵敏、迅速，可用于布南色林原料药中3种遗传毒性杂质的测定。     

乙二醇二甲磺酸酯及其衍生物对抗人绒毛膜促性腺激素的作用

用人绒毛膜促性腺激素(hCG)升高幼大鼠卵巢cAMP含量及增加幼大鼠储精囊重量的方法,试验了乙二醇二甲磺酸酯及其8种衍生物的抗hCG作用。结果表明:乙二醇二甲磺酸酯、甲磺酸氯乙酯及甲磺酸溴乙酯都有明显抗hCG作用;甘油三甲磺酸酯、丙二醇-1,2-二甲磺酸酯则无此作用;甲磺酸乙酯及甲磺酸-β-苯乙酯有抗hCG的储精囊增重作用,但对卵巢cAMP浓度无明显影响;甲磺酸正丙酯及甲磺酸异戊酯在储精囊增重试验中,表现了抗hCG作用。根据以上实验资料,初步分析了乙二醇二甲磺酸酯类抗hCG作用部位及构效关系。     

气相-质谱联用法测定甲磺酸伊马替尼中的甲磺酸烷酯类

目的对甲磺酸伊马替尼中遗传毒性杂质甲磺酸烷酯类进行测定。方法采用气相-质谱法,聚乙二醇PEG-20M(VF-WAXms,30 m×0.32 mm×0.25μm)为固定相的毛细管柱;质谱检测器(MSD),载气为氦气;进样口温度:120℃;柱初始温度40℃,维持3 min;瓶平衡温度:60℃;瓶平衡时间15min;对样品溶液选用Agilent 5977A质谱仪的选择离子检测(SIM)扫描方式进行检测。结果碘甲烷的浓度在1.112~15.56 ng·mL~(-1)内与峰面积线性良好(r=0.9998),方法回收率106.7%;碘代异丙烷的浓度在1.026~14.35 ng·mL~(-1)内与峰面积线性良好(r=0.9998),方法回收率103.2%,稳定性较好。结论该方法快速、灵敏、专属性强,适用于甲磺酸伊马替尼中基因毒性杂质甲磺酸烷酯类的测定。     

棓丙酯注射液热降解动力学及稳定性研究

目的:研究棓丙酯注射液的热降解动力学,为药物贮存提供理论依据。方法:采用高效液相色谱法测定药物含量,并以恒温加速法研究在25、30、40℃下,棓丙酯注射液6月内含量变化及其降解规律,用趋势外推法预测棓丙酯注射液不同温度下的有效期。结果:棓丙酯注射液的降解规律符合一级动力学方程,在25、30、40℃下,其有效期分别为21.7、11.3、5.4个月。结论:温度是影响棓丙酯注射液稳定性的重要因素之一,常温下丙酯注射液是稳定的。     

甲磺酸非诺多泮的化学降解动力学及其制剂的有效期预测

目的：研究甲磺酸非诺多泮的化学降解动力学,来预测制剂的有效期。方法：采用经典恒温法对甲磺酸非诺多泮溶液和甲磺酸非诺多泮注射剂在不同温度不同时间的含量和有关物质进行考察,采用反相高效液相色谱法进行定量分析,通过数学拟合,确定反应动力学方程,求出反应速率常数K。并运用Arrhenius公式,预测甲磺酸非诺多泮溶液和注射剂在常温下的有效期。结果：甲磺酸非诺多泮溶液和注射剂的含量下降符合一级动力学模型,有关物质增加符合零级动力学模型。甲磺酸非诺多泮溶液在25℃时的t0．9为1．09年,其注射剂在25℃时的t0．9为5．12年。结论：通过制剂学手段,显著提高了甲磺酸非诺多泮注射剂的稳定性。     

甲磺酸二氢麦角毒碱溶液鼻黏膜吸收的研究

目的:研究甲磺酸二氢麦角毒碱溶液鼻黏膜吸收规律。方法:考察甲磺酸二氢麦角毒碱在大鼠鼻腔洗出液中的稳定性,在此基础上,以大鼠在体鼻循环为实验模型,研究甲磺酸二氢麦角毒碱溶液的鼻黏膜吸收规律。结果:甲磺酸二氢麦角毒碱在鼻黏膜洗出液中稳定性良好,当循环液体积为5 mL,流速为2．5 mL·min-1时,甲磺酸二氢麦角毒碱溶液鼻黏膜吸收速率常数K不随药物浓度发生变化。结论:甲磺酸二氢麦角毒碱溶液鼻黏膜吸收机制为被动扩散,吸收符合一级动力学,其鼻黏膜平均吸收速率常数K为5．94×10-3min-1。     

Determination of Alkyl Methanesulfonates in Doxazosin Mesylate by Gas Chromatography-mass Spectrometer

High sensitive rapid gas chromatography-mass spectrometry method for the determination of four carcinogenic alkyl methanesulfonates viz. methyl methanesulfonate, ethyl methanesulfonate, isopropyl methanesulfonate and n-butyl methanesulfonate in doxazosin mesylate has been presented by using selective ion monitoring mode. The optimum separation was achieved between methyl methanesulfonate, ethyl methanesulfonate, isopropyl methanesulfonate and n-butyl methanesulfonate on a DB-5 (30 m×0.32 mm×1.0 μm) capillary column under programming temperature. Acetonitrile, water and ammonia (90:9:1 v/v/v) mixture was used as diluent. Various factors involved in the gas chromatography-mass spectrometry method development are also presented. This method was validated as per International Conference on Harmonization guidelines. The limit of quantitation of methyl methanesulfonate, ethyl methanesulfonate, isopropyl methanesulfonate and n-butyl methanesulfonate is 6 ppm with respect to 30 mg/ml of doxazosin mesylate.     

Stability of ertapenem in aqueous solutions

The kinetics of degradation of ertapenem was studied in aqueous solutions at 303, 313, 323 and 333 K and pH 0.42-12.5. Degradation was studied using two methods: HPLC (LiChrospher RP-18 column, 5 microm, 250 mm x 4 mm; mobile phase: methanol-phosphate buffer 25 mmol l(-1), pH 6.5 (15:85, v/v); flow rate--1.2 ml/min; detection UV--298 nm) and UV (294 nm). Specific acid-base catalysis involves: (a) hydrolysis of ertapenem, catalysed by hydrogen ions; (b) hydrolysis of ertapenem dianions catalysed by hydroxide ions; (c) spontaneous hydrolysis of zwitter ions and dianions of ertapenem under the influence of water. The thermodynamic parameters of these reactions--energy, enthalpy and entropy of activation were calculated. It was observed that buffer catalysis occurred in acetate, phosphate and borate buffers.     

Dominant lethal studies with alkylating agents: dose-response relationships.

Dominant lethal studies were conducted to establish dose-response relationships for the mutagenic alkylating agents, methyl methanesulfonate (MMS), and ethyl methanesulfonate (EMS). Male albino mice were treated via single ip injections at doses ranging from 3.125 to 100 mg of MMS/kg and from 50 to 400 mg of EMS/kg. The mutagenic events induced following mating with untreated females were scored on the basis of early fetal deaths. Useful effect dosages were found to be 50 mg/kg for MMS and 200–300 mg/kg for EMS. Dosages of 100 mg of MMS/kg and >300 mg of EMS/kg were judged to be undesirable because of excessive preimplantation losses at these levels. It was not possible to differentiate the response of animals treated with up to 6.25 mg of MMS/kg or 100 mg of EMS/kg from that of the untreated control animals.     

Low level determinations of methyl methanesulfonate and ethyl methanesulfonate impurities in Lopinavir and Ritonavir Active pharmaceutical ingredients by LC/MS/MS using electrospray ionization

Methyl methanesulfonate (MMS) and ethyl methanesulfonate (EMS) have been highlighted as potential genotoxic impurities (PGIs). A sensitive LC/MS/MS method is developed and validated for the determination of MMS and EMS impurities in both Lopinavir and Ritonavir Active pharmaceutical ingredient. Method utilizes, Atlantis T3 column with electrospray ionization in multiple reactions monitoring (MRM) mode for quantitation of impurities. The proposed method is specific, linear, accurate and precise. The calibration curves show good linearity over the concentration range of 0.01-0.23 μg/mL for MMS and 0.005-0.23 μg/mL for EMS. The correlation coefficient obtained is >0.99 in each case. Method has very low limit of detection (LOD) and quantification (LOQ). LOD and LOQ of MMS and EMS are as low as ～0.002 μg/mL and ～0.01 μg/mL respectively. Method has accuracy within 80-120% for both the analytes. This method is a good quality control tool for quantitation of MMS and EMS impurities at very low levels in Lopinavir and Ritonavir.     

Simultaneous determination of alkyl mesilates and alkyl besilates in finished drug products by direct injection GC/MS

We report a specific, fast and feasible method for the simultaneous determination of methyl mesilate (MMS), ethyl mesilate (EMS), isopropyl mesilate (IMS), methyl besilate (MBS) and ethyl besilate (EBS) in finished drug products by GC/MS. Sample preparation was carried out by liquid extraction. The analytes were directly injected into the chromatographic system and quantified with the internal standard method using methyl tosylate (MTS) as internal standard (ISTD). The method gives excellent sensitivity for all the alkyl and aryl esters at typical target analyte level, according to the acceptance criteria that are described in the Guideline on the Limits of Genotoxic Impurities which has been issued in 2007 by the European Medicines Agency (EMA). The average recovery for methanesulfonic acid esters (mesilates) was not lower than 71%, for benzenesulfonic acid esters (besilates) not lower than 94%. A linear range with R2 ≥ 0.9998 has been established for concentrations between 0.01 and 1.33 μg/ml. Validation of the method was carried out on a sample matrix containing MMS, EMS, IMS, MBS and EBS at relevant levels and was further confirmed on finished products containing APIs as mesilate salts (Bromocriptine mesilate, Doxazosin mesilate).     

Development and validation of GC-MS method for the determination of methyl methanesulfonate and ethyl methanesulfonate in imatinib mesylate

A gas chromatography-mass spectrometry (GC-MS) method has been developed for the identification and determination of two carcinogenic and genotoxic mesylate esters viz. methyl methanesulfonate (MMS) and ethyl methanesulfonate (EMS) in imatinib mesylate (INM). The method was optimized based on the peak shapes and resolution of MMS and EMS. The method was validated as per International Conference of Harmonization (ICH) guidelines in terms of limits of detection (LOD), limit of quantitation (LOQ), linearity, precision, accuracy, specificity and robustness. The LOD and LOQ values were found to be 0.3 and 1.0 microg/ml, respectively. The method is linear within the range of 1-15 microg/ml for both the compounds. These mesylate esters were not found in three different batches of pure and pharmaceutical formulations of INM.     

Comparative study of the comet assay and the micronucleus test in amphibian larvae (Xenopus laevis) using benzo(a)pyrene, ethyl methanesulfonate, and methyl methanesulfonate: establishment of a positive control in the amphibian comet assay

The present investigation explored the potential use of the comet assay (CA) as a genotoxicity test in the amphibian Xenopus laevis and compared it with the French standard micronucleus test (MNT). Benzo[a]pyrene (B[a]P), methyl methanesulfonate (MMS), and ethyl methanesulfonate (EMS) were used as model compounds for assessing DNA damage. Damage levels were measured as DNA strand breaks after alkaline electrophoresis of nuclei isolated from larval amphibian erythrocytes using the CA in order to establish a positive control for further ecotoxicological investigations. The results led to the selection of MMS as a positive control on the basis of the higher sensitivity of Xenopus laevis to this compound. The CA and MNT were compared for their ability to detect DNA damage with the doses of chemical agents and exposure times applied. EMS and MMS were shown to increase micronucleus and DNA strand break formation in larval erythrocytes concurrently. However, B[a]P increased micronucleus formation but not that of DNA strand breaks. Time-dose experiments over 12 days of exposure suggest that the CA provides an earlier significant response to genotoxicants than does the MNT. In Xenopus the CA appears to be a sensitive and suitable method for detecting genotoxicity like that caused by EMS and MMS. It can be considered a genotoxicity-screening tool. The results for B[a]P show that both tests should be used in a complementary manner on Xenopus.     

Toxicity of Four Alkylating Agents on in Vitro Rat Embryo Differentiation and Development

The relative developmental toxicity of four direct acting, alkylatingagents was determined in primary cultures of differentiatingrat embryo midbrain (CNS) and limb bud (LB) cells and comparedwith that observed in the rat whole embryo postimplantationculture system. The alkylating agents tested include methylnitrosourea(MNU), ethylnitrosourea (ENU), methyl methanesulfonate (MMS),and ethyl methanesulfonate (EMS). These alkylating agents havebeen shown to produce developmental toxicity following eitherin vitro or in vivo exposure. Viability for both CNS and LBwas assessed by a neutral red dye assay. Differentiation ofCNS cells was assessed by hematoxylin staining of neurons; differentiationof LB cells was assessed by Alcian blue staining of extracellularproteoglycans. Relative potencies of these compounds in thecell culture system were not the same as those observed in theembryo culture system. Whereas rank order of potency in thecell culture system, for viability and differentiation, wasMMS > MNU > ENU > EMS, rank order in the embryo culturesystem, for embryo lethality and malformations, was MNU >ENU > MMS > EMS. Effective concentrations for cell cultureviability and differentiation by MNU and ENU in cell culturewere about three to nine times higher than comparable valuespreviously reported for embryos, while effective concentrationsfor MMS and EMS were two to seven times lower than those observedin the embryos. Differences in potency between the two culturesystems may be related to differences in formation and repairof DNA adducts, as well as differences in culture conditions.     

2-Chloroethyl (methylsulfonyl)methanesulfonate and related (methylsulfonyl)methanesulfonates. Antineoplastic activity in vivo

2-Haloethyl and ethyl (methylsulfonyl)methanesulfonates were prepared via sulfene intermediates. 2-Chloroethyl (methylsulfonyl)methanesulfonate is highly active against P388 leukemia in vivo; the majority of leukemic mice treated with this compound at 50 mg/kg per day, qd 1-5, survived more than 30 days and about 37% survived for more than 60 days. 2- Fluoroethyl (methylsulfonyl)methanesulfonate is also highly effective against P388 cells in vivo, but it is more toxic. Other (methylsulfonyl)methanesulfonate esters are more active than the analogous methanesulfonates and chloromethanesulfonates .     

EMS in Viracept—Initial (‘traditional’) assessment of risk to patients based on linear dose response relations

Prior to having performed in depth toxicological, genotoxicological and DMPK studies on ethyl methanesulfonate (EMS) providing solid evidence for a thresholded dose response relationship, we had prepared and shared with regulatory authorities a preliminary risk estimate based on standard linear dose–effect projections. We estimated that maximal lifetime cancer risk was in the order of 10⁻³ (for lifetime ingestion of the maximally contaminated tablets) or 10⁻⁴ for the exposure lasting for 3 months. This estimate was based on a lifetime cancer study with methyl methanesulfonate (MMS; as insufficient data were available for EMS) in rodents and default linear back extrapolation. Analogous estimates were made specifically for breast cancer based on short term tumorigenicity studies with EMS in rats, for the induction of heritable mutations based on specific locus and dominant lethal tests in mice and for the induction of birth defects based on teratogenicity studies in mice. We concluded that even under worst case assumptions of linear dose relations the chance of experiencing these adverse effects would be very small, comprising at most a minute additional burden among the background incidence of the patients.