Chemical derivatization as a strategy to enhance detectability of agents used in cancer management

The bioanalysis of drugs used in the management of cancer is often complicated by the lack of selectivity and sensitivity. Chemical derivatization of these drugs prior to their chromatographic analysis represents a viable strategy to improve chromatographic resolution and to enhance detectability. This review provides examples of how this approach can meet these objectives. Derivatization of racemic cyclophosphamide with a chiral acylating agent, following hydroxyalkylation to introduce a reactive centre into the molecule, provides the basis for its stereospecific analysis. The analysis of dianhydrogalactitol is described, in which diethyldithiocarbamate is used as a nucleophilic derivatizing agent that improves chromatographic behaviour and analytical sensitivity. The final example that is described is the design and preparation of improved fluorogenic reagents (o-phthalaldehyde analogues) for the derivatization of peptides and application of these reagents to the trace analysis of leu-enkephalin in plasma.     

盐酸芬氟拉明对映体的柱前衍生物GC法分析

以ω－莰烷酰氯为衍生试剂,采用柱前衍生化ＧＣ法,成功地分析了盐酸芬氟拉明、金刚乙胺和盐酸氯胺酮。光谱分析确证了芬氟拉明酰胺衍生物的结构,优化了盐酸芬氟拉明酰胺衍生物的结构,优化了盐酸芬氟拉明衍生条件,建立了盐酸花氟拉明的内标法定量分析方法。     

用均匀设计法优化β—环糊精手性固定相的衍生化反应工？…

采用均匀设计法对衍生化β－环糊精（β－ＣＤ）键合固定相的合成条件进行优化,考察的主要因素包括衍生化试剂异氰酸苯酯的用量、反应时间及反应温度。衍生化后β－ＣＤ键合相的碳含量优化结果预测值和实测值分别为１７．９５％和１８．１０％。采用红外光谱、热重分析曲线对衍生化前后β－ＣＤ键合固定相进行表征,探讨了衍生化前后β－ＣＤ键合相的手性拆分能力。     

清开灵注射液生产中板蓝根提取液氨基酸分析

目的分析清开灵注射液生产过程中所用板蓝根提取液的氨基酸的成分。方法柱前衍生化反相高效液相色谱法,色谱柱为ZorbaxEclipse氨基酸分析柱,邻苯二甲醛（OPA）和9-芴甲基氯甲酸酯（FMOC）使氨基酸在柱前发生衍生化以用于色谱分析。结果板蓝根提取液中至少含有15种氨基酸,其中精氨酸、脯氨酸、苏氨酸、缬氨酸和丙氨酸的含量最高,板蓝根提取液是清开灵注射液中精氨酸的主要来源。结论为强化清开灵注射液生产的过程控制和提高产品的质量标准提供了研究思路。     

柱前衍生化HPLC分析蒲公英多糖的单糖组成

目的 合理优化衍生方法测定蒲公英多糖中单糖的组成及摩尔比。方法 采用1-苯基-3-甲基-5-吡唑啉酮(PMP)柱前衍生,用反相高效液相色谱法进行测定。色谱条件为：ZORBAX-C₁₈柱;流动相：0.1 mol·L^-1的磷酸盐缓冲液(pH 6.7)-乙腈(83∶17);流速：1 mL·min^-1;检测波长：245 nm;进样量：20 μL;柱温：室温。结果 蒲公英多糖由D-鼠李糖、葡萄糖、D-半乳糖、D-木糖和D-阿拉伯糖组成,以不同方法提取的各单糖含量的摩尔比为：水提取法,Rha∶Glc∶Gal∶ Ara=0.088∶0.500∶0.190∶0.340;超声提取法,Rha∶Glc∶Gal∶Ara=0.050∶0.350∶0.080∶0.320;酶提取法,Rha∶Glc∶ Gal∶Ara=0.270∶0.630∶0.330∶0.460;超声协提取同酶法,Rha∶Glc∶Gal∶Ara =0.078∶0.550∶0.130∶0.170。结论 改进后的衍生化方法和等度洗脱的色谱方法具有操作简单、快速、重复性好等特点,可用于蒲公英多糖中单糖组成和摩尔比的测定。     

Recent advances in trace analysis of pharmaceutical genotoxic impurities

Genotoxic impurities (GTIs) in pharmaceuticals at trace levels are of increasing concerns to both pharmaceutical industries and regulatory agencies due to their potentials for human carcinogenesis. Determination of these impurities at ppm levels requires highly sensitive analytical methodologies, which poses tremendous challenges on analytical communities in pharmaceutical R&D. Practical guidance with respect to the analytical determination of diverse classes of GTIs is currently lacking in the literature. This article provides an industrial perspective with regard to the analysis of various structural classes of GTIs that are commonly encountered during chemical development. The recent literatures will be reviewed, and several practical approaches for enhancing analyte detectability developed in recent years will be highlighted. As such, this article is organized into the following main sections: (1) trace analysis toolbox including sample introduction, separation, and detection techniques, as well as several ‘general’ approaches for enhancing detectability; (2) method development: chemical structure and property-based approaches; (3) method validation considerations; and (4) testing and control strategies in process chemistry. The general approaches for enhancing detection sensitivity to be discussed include chemical derivatization, ‘matrix deactivation’, and ‘coordination ion spray-mass spectrometry’. Leveraging the use of these general approaches in method development greatly facilitates the analysis of poorly detectable or unstable/reactive GTIs. It is the authors’ intent to provide a contemporary perspective on method development and validation that can guide analytical scientists in the pharmaceutical industries.     

A novel pre-column fluorescent derivatization method for the sensitive determination of aristolochic acids in medicinal herbs by high-performance liquid chromatography with fluorescence detection

Aristolochic acids (AAs) are a family of structurally related nitrophenanthrene carboxylic acids existing in Aristolochia, Bragantia, and Asarum species. AAs have been proven to have nephrotoxic and carcinogenic toxicity. In this study, a novel pre-column fluorescence derivatization procedure followed by high-performance liquid chromatography-fluorescence detection (HPLC-FLD) is developed for the analysis of AAs in medicinal herbs. The nitro group in the phenanthrene ring of AAs was removed by NaBH₄ in water–THF (2:1, v/v), resulting in the corresponding aristolic acids. The analysis of AAs in medicinal herbs was based of the sensitive fluorescence detection of aristolic acids after the chemical derivatization. Because the produced aristolic acids are highly fluorescent the limit of detection (LOD) of AAI and AAII were lowered to 0.06 and 0.04 ng/mL, respectively, which is at least an order of magnitude lower than those in the reported HPLC and LC–MS methods. Good linearity with correlation coefficients higher than 0.997 were obtained for AAI and AII in the calibration ranges of 0.2–800 ng/mL. The derivatization conditions such as reaction temperature, time and the amount of NaBH₄ were optimized. The developed method provided satisfactory intra-day and inter-day precisions with RSDs less than 1.4% and 3.8%, respectively. The relative analytical error was less than 7% for the analysis of AAI and AAII in spiked matrix samples.     

柱前衍生化-HPLC法测定患者体内丙戊酸钠的血药浓度

目的建立反相高效液相色谱法测定人血清中丙戊酸钠血药浓度。方法血清用环己烷提取,以环已烷羧酸为内标,2-溴苯乙酮为衍生化试剂,用高效液相色谱法测定丙戊酸钠血药浓度。采用Symmetry C18(4.6 mm×250 mm,5μm)色谱柱;流动相为甲醇-水(82∶18);检测波长为248 nm;流速为1.0 ml.min-1;柱温:30℃。结果血清中丙戊酸钠线性范围为12.5～150mg.L-1,平均回收率98.74%,日内RSD<5%。结论该法快速、灵敏、准确,适用于临床常规监测需要。     

免疫亲和柱净化光化学衍生高效液相色谱-荧光检测法测定动物类药材中黄曲霉毒素

目的建立动物类药材中黄曲霉毒素B1,B2,G1,G2的免疫亲和柱净化光化学衍生高效液相色谱-荧光检测(HPLC-FLD)法。方法样品经甲醇-水(80∶20)提取后,通过免疫亲和柱净化、柱后光化学衍生、高效液相色谱-荧光检测器测定。结果在优化条件下,黄曲霉毒素G2,G1,B2,B1的检出限分别为0.15,0.25,0.1,0.2μg/kg,回收率为78.8%～106.7%,RSD均低于7.1%。结论所用方法简便快速、灵敏度高、重现性好,可满足动物类药材中黄曲霉毒素检测的需要。     

邻苯二甲醛-9-氯甲酸芴甲酯柱前衍生HPLC法测定血必净注射液中氨基酸

目的通过建立HPLC分析法测定血必净注射液中氨基酸,为提高中药注射液质量标准提供依据.方法通过OPA-FMOC柱前衍生化法,建立同时检测血必净注射液中18种常见氨基酸的液相色谱法.色谱柱为Agilent SB-C18柱(250mm×4.6 mm,5 μm);流动相A为0.02 mol/L醋酸钠溶液(含200μL/L三乙...