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??OBJECTIVE To compare the HPLC-PAD method and HPLC-ELSD method for determination of related substances in spectinomycin hydrochloride for injection, and determine 31 batches of samples by the two methods. METHODS The following conditions were used for both methods:the analytical column was Apollo C₁₈ column (4.6 mm??250 mm,5 ??m), mobile phase was 0.1 mol??L^-1 trifluoroacetic acid solution, the column temperature was maintained at 30 ??, and the sample volume was 20 ??L. For the PAD detection, gold electrode (3 mm)was used as the detection electrode, Ag-AgCl as reference electrode, titanium alloy as counter electrode, and integrated amperometric detector was employed with four waveform detection potential. For the ELSD detection, the drift tube temperature was set at 110 ??, and the carrier gas flow rate was 2.6 L??min^-1, with gain of 1. RESULTS The two methods were consistent in the species and number of detected impurities. The detection limits of the PAD method and ELSD method were 2.4 and 72.8 ng, respectively (S/N=3). For the PAD method there existed a direct linear relationship in the concentration range of 0.15-150 ??g??mL^-1, while a double logarithmic linear relationship was shown in the concentration range of 17-170 ??g??mL^-1 for the ELSD method. When 31 batches of samples were determined for related substances by the two kinds of methods, the contents of impurity D and E were basically the same, but there were significant differences in the contents of impurity A and (4R)-dihydrospectinomycin. After correction, the contents of impurity A and total impurities determined by the PAD method were consistent with those by the ELSD method. CONCLUSION The two methods can both effectively detect the related substances of spectinomycin to achieve the purpose of controlling related substances. When using HPLC-PAD method, the impurity A and (4R)-dihydrospectinomycin should be adjusted by using response factor or the reference substances. The ELSD method needs to use double log linear regression.     

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??OBJECTIVE To identify the related substances in aituomode bulk drug. METHODS HPLC-QTrap-MS and HPLC-QTOF-MS were used to determine seven kinds of related substances denoted by Imp-A to Imp-G. RESULTS The structures and molecular weights of the related substances in aituomode were identified. CONCLUSION The method is simple and accurate, providing a good idea for the identification of the related substances in bulk drugs. The experimental data are valuable to the determination of the related substances and quality control of aituomode.     

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??OBJECTIVE To establish an RP-HPLC method for determination of the contents of eplerenone and its related substances. METHODS A RP-HPLC method was developed by using a C₁₈ column (Agilent Poroshell,4.6 mm??100 mm, 2.7 ??m), the mobile phase consisted of H₂O-CH₃CN-MeOH (64??18??18), the flow rate was 1.0 mL??min^-1, and the detection wavelength was set at 240 nm. The diluent was H₂O-CH₃CN-MeOH (50??25??25). RESULTS Eplerenone was well separated from the impurities. For the determination of related substances, the LODs of eplerenone, its intermediate (?? and ??) and related substances (A-E) were 0.03, 0.13, 0.25, 0.05, 0.05, 0.26, 0.07 and 0.12 ng, and the LOQs were 0.12, 0.42, 0.84, 0.18, 0.18, 0.86, 0.22 and 0.41 ng, respectively. The calibration curves of eplerenone and related substances (A-E) had good linear relationship within 0.13-1.53, 0.25-3.03, 0.25-2.99, 0.13-1.61, 0.14-1.63, and 0.13-1.54 ??g??mL^-1(r=0.999 9, n=7), and the correction factors of the related substances (A-E) were all in the range of 0.90-1.10. The average recoveries of the related substances (A-E) were all within 95.0%-105.0% (n=9). For the content determination of eplerenone, the validation test showed good linearity over the range of 12.60-75.63 ??g??mL^-1 (r=0.999 6, n=6) with good repeatability (RSD=0.19%, n=6). CONCLUSION The developed method proves to be simple, accurate, specific and reliable. It can be applied to the determination of eplerenone and its related substances.     

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??OBJECTIVE To establish a UPLC-MS method for the separation and detection of principal component isomers and related substances in ioversol bulk drug. METHODS The separation was performed on a Waters ACQUITY UPLC^TM BEH C₈ column (2.1 mm??100 mm,1.7 ??m), the mobile phase consisted of acetonitrile-0.1% formic acid in water (3:97),and the detection wavelength was set at 254 nm. The identification of isomers and impurities in ioversol bulk drug was performed with a triple-quadrupole mass spectrometer, with an electrospray ionization (ESI) source in the positive ion mode, and the cone voltage was set at 80 V. RESULTS The four optical isomers and related substances were seperated well, and their structures were confirmed by MS,UV, and RP-UPLC.The linear ranges of ioversol, impurity??, and impurity ?? were 0.5-52.8, 0.2-9.3, and 0.5-28.4 ??g??mL^-1, respectively (r=0.999 9); the detection limits of ioversol, impurity??, and impurity?? were 0.2, 0.08, and 0.2 ??g??mL^-1, respectively.CONCLUSION The method is simple, sensitive, and accurate, can be used to separate and determine the principal component isomers and related substances in ioversol bulk drug.     

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??OBJECTIVE To establish an improved reversed-phase high performance liquid chromatography method coupled with pulsed electrochemical detection for determining the related substances of netilmicin sulfate injection. METHODS Agilent Proshell 120 SB-C₁₈ column (4.6 mm×150 mm,2.7 μm)and gradient elution were used. Mobile phase A was 0.2 mol·L^-1 trifluoroacetic acid in 0.1 mol·L^-1 sodium hydroxide solution-acetonitrile (97:3), mobile phase B was 0.1% pentafluoropropionic acid-acetonitrile (97:3), and the flow rate was 0.8 mL·min^-1. A pulsed electrochemical detector was adopted, and the temperatures of detector and column were kept at 35 ??. The working electrode was a gold electrode with diameter of 3 mm and a quadruple-potential waveform (QPW)was selected as detection waveform. The injection volume was 25 μL. NaOH solution of 0.8 mol·L^-1 was added post-column at a flow rate of 0.3 mL·min^-1. RESULTS A total of 28 impurities could be detected and effective separation was achieved in the typical sample and most of which could not be separated in the method of Ch.P 2015. The linearity of the calibration curve for netilmicin ranged from 0.25 to 15 μg·mL^-1 with a coefficient of determination equal to 0.999 1. The LOD and LOQ of netilmicin were found to be 0.25 ng and 1.25 ng, respectively. The repeatability RSD(n=6) of the single largest impurity and total impurities were 0.9% and 0.8%, respectively. The sample solution was stable within 24 h. CONCLUSION Compared with previously published investigations, the improved method shows higher sensitivity, better separation ability and good reproducibility, especially for differentiating the origin of bulk drug for netilmicin sulfate injection, thus is more suitable for the determination of related substances of netilmicin sulfate injection.     

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??OBJECTIVE To investigate the sustained-release ability of Na-montmorillonite (Na-MMT) on 1-deoxynojirimycin (DNJ) in mulberry leaf extract (MLE). METHODS In order to prepare DNJ-MMT composite, DNJ was intercalated into the interlayer of MMT by stirring. Time and temperature for adsorption and ratio of DNJ to MMT were determined by calculating the loading of DNJ, then in vitro release experiment was carried out to estimate the sustained-release characteristics of DNJ-MMT. RESULTS DNJ-MMT composite with a drug loading of 111.64 mg??g^-1 was obtained by stirring 5.00 g Na-MMT and 16.00 g MLE containing 4 016.0 mg DNJ in 1 000 mL deionized water at 100 ?? for 1 h. The composite released DNJ quickly in water, potassium phosphate buffer (pH 7.4), and 0.1 mol??L^-1 hydrochloric acid solution. With the increasing of volume of hydrochloric acid solution, the release ratio of DNJ also increased correspondingly. CONCLUSION The sustained-release effect of Na-MMT on DNJ is proved. Na-MMT can be used for study of DNJ sustained-release preparations.     

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??OBJECTIVE To establish an HPLC method to determine the related substances of vortioxetine hydrobromide and identify the degradation products of vortioxetine hydrobromide by HPLC-MS. METHODS An HPLC method was developed by using a CN column(Agilent Zorbax SB-CN,4.6 mm??250 mm, 5 ??m),the mobile phase consisted of CH₃CN-HCOONH₄(50??50, pH adjusted to 5.0 with phosphoric acid),the flow rate was 1.0 mL??min^-1, and the detection wavelength was set at 228 nm. The column temperature was maintained at 25 ??, and the injection volume was 20 ??L. RESULTS Vortioxetine hydrobromide was well separated from the related substances. The detection sensitivity of vortioxetine hydrobromide and its related substances met the determination requirements. The calibration curves of vortioxetine hydrobromide and related substances had good linearity. The repeatability of the method was good(RSD=6.89%, n=6). The average recoveries of the sample were all within 95%-105%. The degradation product produced by oxidation was identified by mass spectrometry as related substance D. CONCLUSION The developed method proves to be simple,accurate,specific and reliable. It can be applied to the determination of vortioxetine hydrobromide and its related substances.     

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??OBJECTIVE To prepare cholesterol-PEG-modified amphotericin B liposomes and amphotericin B liposomes, and evaluate their encapsulation efficiency and pharmacokinetics. METHODS It was employed to prepare both kinds of liposomes with the film-evaporation, the dynamic light scattering techniques and Ultrafree-CL were used to determine the mean diameter and the encapsulation efficiency, respectively. Pharmacokinetics was evaluated in Wista rats. RESULTS Mean diameter of the liposomes was (115??20) nm,and the encapsulation efficiency was over 99.9%. Moreover, the pharmacokinetic study in rats indicated that cholesterol-PEG-modified liposomes could significantly increase the ??_max and AUC. CONCLUSION Amphotericin B liposomes prepared by film-evaporation method have high encapsulation efficiency, and cholesterol-PEG modification can prolong the circulation time of amphotericin B in vivo.     

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??OBJECTIVE To establish an HPLC-HRMS/MS method for analyzing the structures and sources of the related substances in etimicin sulfate. METHODS The chromatographic separation was achieved on a broad pH range column with a basic elution system composed of[H₂O-ammonia-glacial acid(96??3.6??0.4)]-methanol(70??30). Twenty percent of the eluent was detected under positive electrospray ionization(ESI) by Q-Exactive mass spectrometry. The fragmentation pathways were elucidated according to the HRMS and HRMS/MS fragmentation of etimicin and some known compounds. Then the unknown related substances were identified by analyzing their HRMS and HRMS/MS fragmentation with the help of the rule. RESULTS Sixty-five related substances were detected by the HPLC-HRMS/MS method in the two samples from different companies,among which 38 were detected in the product of company A, 59 in that of company B, and 32 were detected for both companies. Fifty-four related substances were identified or deduced, and the other 11 were not able to be identified due to limited information. Based on the significant difference of impurity spectra between the two enterprises, the key parameters of the synthesis process were analyzed. CONCLUSION An HPLC-HRMS/MS method, which showes excellent accuracy, is developed to identify the related substances in etimicin sulfate. The resources and structures of the related substances are analyzed, which will be helpful to the process optimization.     

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??OBJECTIVE To construct a novel daunorubicin-loaded microparticles (DNR-MPs)drug delivery system, and study its proliferation inhibiton effect on leukemia cells. METHODS Both DNR-MPs-IDL(DNR microparticles with intracellular drug loading method)and DNR-MPs-EDL (DNR microparticles with extracellular drug loading method)were prepared from HL-60 cells, and the average particle size, Zeta potential and drug loading efficiency of DNR-MPs were measured. The uptake of DNR-MPs by HL-60 cells was analyzed by confocal laser scanning microscopy (CLSM)and flow cytometry (FCM). The inhibition effect of DNR-MPs on the proliferation of HL-60 cells was evaluated by MTT assay. RESULTS There was no significant difference in the average particle size and Zeta potential of the two DNR-MPs. The drug loading efficiency of DNR-MPs-EDL was higher compared with DNR-MPs-IDL. The uptake RESULTS showed that the two DNR-MPs significantly increased the drug uptake compared with free DNR (P<0.05). DNR-MPs-IDL showed a higher drug uptake by the cells than DNR-MPs-EDL did (P<0.05). They also exhibited an enhanced inhibition effect on the proliferation of HL-60 cells compared with DNR (P<0.05). CONCLUSION The different ways of preparation of drug-loaded microparticles can cause varying anti-tumor effect. Microparticles can significantly augment the anti-leukemia efficacy of DNR, indicating a promising drug carrier for the therapy of leukemia.     

高效液相色谱法测定氨甲环酸注射液的含量和有关物质

 目的 建立反相高效液相色谱法,测定氨甲环酸注射液的含量,并考察其有关物质。方法 以C₁₈柱为固定相;以0.23% 十二烷基硫酸钠溶液（取磷酸二氢钠18.3 g,加水800 mL溶解,加三乙胺8.3 mL混匀后,再加入2.3 g十二烷基硫酸钠,溶解混匀,用磷酸调节pH至2.5,加水至1 000 mL）-甲醇（60∶40）为流动相;检测波长为220 nm。结果 氨甲环酸在0.54～5.38 g·L-1内线性关系良好,相关系数0.999 9,最低检测限75 ng,低、中、高3种浓度的平均回收率分别为99.8%,100.4%,100.1%。结论 本方法准确、简便,专属性强,可用于氨甲环酸注射液的含量测定和有关物质的考察。     

RP-HPLC测定依折麦布原料药中的有关物质

目的: 建立测定依折麦布原料药中有关物质的RP-HPLC测定法。方法: Phenomenex Luna C₁₈色谱柱(4.6 mm ×150 mm,5 μm),流动相0.01%磷酸(A),乙腈(B),梯度洗脱,流速1.0 mL·min^-1,检测波长231 nm,柱温35℃。结果: 依折麦布与有关物质分离良好;有关物质A,B,C在0.035~0.8 mg·L^-1峰面积与浓度呈良好的线性关系,r分别为0.999 7,0.999 4,0.999 9(n=6)。结论: 方法操作简便,灵敏度高,专属性强,重复性好,可用于依折麦布原料药中有关物质的检查。     

RP-HPLC测定阿折地平含量及有关物质

阿折地平（azelnidipine）是一种新型的长效钙拮抗剂，可以选择性的阻滞L型钙通道。阿折地平由日本三共制药开发，2003年在日本上市，用于高血压的治疗。关于阿折地平的分析方法笔者尚未见国内报道。本实验建立了用反相高效液相色谱法测定阿折地平含量及有关物质的方法，可用于阿折地平原料药的质量控制。     

HPLC测定长春西汀注射液的有关物质

目的:应用高效液相色谱法测定长春西汀注射液的有关物质.方法:采用Hypersil ODS C18色谱柱(4.6 mm×150 mm,5μm),甲醇-1·75 g·L-1碳酸铵溶液-乙酸(80∶25∶3)为流动相,流速1.0 mL· min-1,检测波长273 nm.结果:在选定的色谱条件下,长春西汀与有关物质分离完全,最低检测限为20 ng,精密度良好(RSD 0.52％).结论:方法简便、准确、专属性好、灵敏度高,可用于长春西汀注射液的有关物质测定.     

复方甘草酸苷中有关物质的测定

目的 建立同时测定复方甘草酸苷片中3个主成分有关物质的反相高效液相色谱法(二极管阵列检测器),以控制其质量.方法采用C18色谱柱,以0.025 mol·L-1醋酸钠溶液(pH 3.5)-乙腈(59:41)洗脱,检测波长251,360 nm. 结果 样品中各主成分与有关物质的分离度及检出灵敏度可满足测定要求,甘草酸单铵盐,甘氨酸,DL-蛋氨酸最低检出限分别为2.02,0.28,0.96 ng.结论该法灵敏、准确、简便、专属性强,可作为该制剂质量控制的有效手段.     

RP-HPLC测定舒他兰锌及其有关物质

 目的建立反相高效液相色谱法测定抗癌光敏剂舒他兰锌(Suftalan zinc)含量及有关物质。方法采用SHIMADZU SHIM-PACK VP-ODS色谱柱(4.6 mm×150 mm,5μm),以四氢呋喃-甲醇-N,N二甲基甲酰胺-水-三乙胺(150∶105∶755∶1 000∶1.4,磷酸调pH值6.5～7.0)为流动相,流速1.0 mL·min^-1,紫外可见检测器于677 nm测定。结果RP-HPLC测定的线性范围为20.0～120.1 mg·L^-1,相关系数r=0.999 9;最低检测限为0.048μg;样品溶液至少在3 d内稳定;日内精密度(RSD=0.70%)良好。结论采用反相高效液相色谱法测定抗癌光敏剂(舒他兰锌)含量及有关物质方法简便,结果准确。     

RP-HPLC测定染料木素-4''''-葡萄糖苷及其有关物质

目的 建立染料木素-4'-葡萄糖苷的含量测定方法.方法 采用反相高效液相色谱法.Diamonsil C18柱(4.6 mm×250 mm,5 μm),流动相为乙腈-水(用磷酸调节pH至2.73)(2575),流速1.0 mL·min-1,检测波长261 nm,柱温35℃.结果 染料木素-4'-葡萄糖苷在0.505 5～101.1 mg·L-1内峰面积与浓度呈良好线性关系(r=1.000 0,n=6),平均回收率99.2%(RSD=1.1%,n=9),日内精密度RSD＜0.5%,日间精密度RSD＜0.7%,最低检测限0.2 ng.同时对样品中所含染料木素、染料木素-7-葡萄糖苷和染料木素-7,4'-二葡萄糖苷等微量有关物质的分离和检测也获得满意的结果.结论 该方法简便、可靠,可用于染料木素及其苷类的质量控制.     

梯度洗脱高效液相色谱法检测他克莫司有关物质

 目的建立他克莫司有关物质检测的方法,并观察样品在不同溶剂中可能存在的异构体的转化情况,同时液-质联用技术进一步确证有关物质。方法以辛烷基硅烷键合硅胶为填充剂,以0.02mol·L^-1磷酸二氢钾(pH3.5)与乙腈为流动相梯度洗脱;检测波长为210nm,柱温60℃;利用液-质联用技术确证可能存在的杂质。结果样品在质子性的溶剂中确实存在异构体的转化且该转化能达到平衡,但在乙腈溶剂中基本无转化,该溶剂可作为他克莫司有关物质的测定溶剂,样品中的主要杂质为发酵副产物。结论本法简便、准确,重复性好,能有效地检测样品中的有关物质。     

HPLC测定维生素C注射液有关物质含量

 目的 建立以HPLC测定维生素C注射液中有关物质的含量。方法 采用菲罗门公司NH₂色谱柱(4.6 mm ×250 mm,5 μm),柱温：45 ℃;以磷酸盐缓冲液(6.8 g磷酸二氢钾用水溶解并稀释至1 000 mL)-乙腈(30∶70)为流动相;流速：1.0 mL·min-1;检测波长：210 nm。结果 该方法定量限为45 ng,破坏出的杂质峰能达到基线分离,抗氧剂不干扰杂质测定。结论 该方法灵敏度高、专属性强、准确,适用于维生素C注射液中有关物质的检测。